Search

The homozygous disruption of the mouse AP-2 gene yields a complex and lethal phenotype that results from defective development of the neural tube, head, and body wall. The severe and pleiotropic developmental abnormalities observed in the knockout mouse suggested that AP-2 may regulate several morphogenic pathways. To uncouple the individual developmental mechanisms that are dependent on AP-2, we have now analyzed chimeric mice composed of both wild-type and AP-2-null cells. The phenotypes obtained from these chimeras indicate that there is an independent requirement for AP-2 in the formation of the neural tube, body wall, and craniofacial skeleton. In addition, these studies reveal that AP-2 exerts a major influence on eye formation, which is a critical new role for AP-2 that was masked previously in the knockout mice. Furthermore, we also have uncovered an unexpected influence of AP-2 on limb pattern formation; this influence is typified by major limb duplications. The range of phenotypes observed in the chimeras displays a significant overlap with those caused by teratogenic levels of retinoic acid, strongly suggesting that AP-2 is an important component of the mechanism of action of this morphogen.

International audience; The ecotropic viral integration site 1 (Evi1) oncogenic transcription factor is one of a number of alternative transcripts encoded by the Mds1 and Evi1 complex locus (Mecom). Overexpression of Evi1 has been observed in a number of myeloid disorders and is associated with poor patient survival. It is also amplified and/or overexpressed in many epithelial cancers including nasopharyngeal carcinoma, ovarian carcinoma, ependymomas, and lung and colorectal cancers. Two murine knockout models have also demonstrated Evi1's critical role in the maintenance of hematopoietic stem cell renewal with its absence resulting in the death of mutant embryos due to hematopoietic failure. Here we characterize a novel mouse model (designated Evi1fl3) in which Evi1 exon 3, which carries the ATG start, is flanked by loxP sites. Unexpectedly, we found that germline deletion of exon3 produces a hypomorphic allele due to the use of an alternative ATG start site located in exon 4, resulting in a minor Evi1 N-terminal truncation and a block in expression of the Mds1-Evi1 fusion transcript. Evi1δex3/δex3 mutant embryos showed only a mild non-lethal hematopoietic phenotype and bone marrow failure was only observed in adult Vav-iCre/+, Evi1fl3/fl3 mice in which exon 3 was specifically deleted in the hematopoietic system. Evi1δex3/δex3 knockout pups are born in normal numbers but die during the perinatal period from congenital heart defects. Database searches identified 143 genes with similar mutant heart phenotypes as those observed in Evi1δex3/δex3 mutant pups. Interestingly, 42 of these congenital heart defect genes contain known Evi1-binding sites, and expression of 18 of these genes are also effected by Evi1 siRNA knockdown. These results show a potential functional involvement of Evi1 target genes in heart development and indicate that Evi1 is part of a transcriptional program that regulates cardiac development in addition to the development of blood.

Thirty percent of human breast cancers have amplification of ERBB2, often in conjunction with mutations in p53. The most common p53 mutation in human breast cancers is an Arg-to-His mutation at codon 175, an allele that functions in a dominant oncogenic manner in tumorigenesis assays and is thus distinct from loss of p53. Transgenic mice expressing mouse mammary tumor virus-driven neu transgene (MMTV-neu) develop clonal mammary tumors with a latency of 234 days, suggesting that other events are necessary for tumor development. We have examined the role of mutations in p53 in tumor development in these mice. We have found that 37% of tumors arising in these mice have a missense mutations in p53. We have directly tested for cooperativity between neu and mutant p53 in mammary tumorigenesis by creating bitransgenic mice carrying MMTV-neu and 172Arg-to-His p53 mutant (p53-172H). In these bitransgenic mice, tumor latency is shortened to 154 days, indicating strong cooperativity. None of the nontransgenic mice or the p53-172H transgenic mice developed tumors within this time period. Tumors arising in the p53-172H/neu bitransgenic mice were anaplastic and aneuploid and exhibited increased apoptosis, in distinction to tumors arising in p53-null mice, in which apoptosis is diminished. Further experiments address potential mechanisms of cooperativity between the two transgenes. In these bitransgenic mice, we have recapitulated two common genetic lesions that occur in human breast cancer and have shown that p53 mutation is an important cooperating event in neu-mediated oncogenesis.

Neuroptera Linnaeus, 1758, Published as part of Shi, Chaofan, Makarkin, Vladimir N., Yang, Qiang, Archibald, S. Bruce & Ren, Dong, 2013, New species of Nymphites Haase (Neuroptera: Nymphidae) from the Middle Jurassic of China, with a redescription of the type species of the genus, pp. 393-410 in Zootaxa 3700 (3) on page 395, DOI: 10.11646/zootaxa.3700.3.4, http://zenodo.org/record/248765

Nymphites priscus (Weyenbergh, 1869) (Figs 1, 2) Insecte: Winkler 1865: 429 (listed); Winkler 1896: 314 (listed). Hemerobius priscus Weyenbergh 1869 a: 264; Pl. 34, Figs 13, 14 (original description); Weyenbergh 1869 b: 236 (listed); Weyenbergh 1874: 97 (listed); Oppenheim 1888: 226, 227 (as a synonym of Chrysopa excelsa Hagen, 1862 [= Osmylites excelsa Oppenheim, 1888]); Scudder 1891: 148 (listed); Martins-Neto & Vulcano 1989: 369, 370 (as a synonym of Nymphites braueri Haase, 1890). Névroptère: Meunier 1897: 238 (notes). Nymphites priscus: Haase 1890: 23 (taxonomic notes); Handlirsch 1906–1908 [1906]: 609 (taxonomic notes); Martynova 1949: 167 (listed); Lambkin 1988: 450, 453 (mentioned); Martins-Neto 1992: 120 (mentioned); Makarkin & Archibald 2003: 175 (taxonomic notes); Ponomarenko 2003: 90 (mentioned); Makarkin & Menon 2005: 810 (taxonomic notes). Diagnosis. Distinguished from Nymphites bimaculatus sp. nov. and from Nymphites sp. A by the absence of crossveins between distal branches of CuA and branches of MP. Description. Thorax fragmentarily preserved; details not discernible. Hind wing approximately 29 mm long as preserved (estimated complete length 31 mm); 8.0 mm wide as preserved (estimated complete width 8.5 mm). Margins poorly-preserved; trichosors not visible. Costal space narrow. Subcostal veinlets simple in proximal 2 / 3, some distal veinlets apparently shallowly forked. ScP, RA preserved as very closely approaching veins; subcostal space not visible. Preserved veinlets of ScP + RA dichotomously branched. RP smooth, only slightly zigzagged; originating near wing base. Preserved crossveins between RA, RP before fusion of ScP + RA widely spaced, rather regularly arranged. RP with 11 or 12 branches (poorly preserved); at least RP 1, RP 2 deeply forked. Crossveins in radial space relatively scarce, irregularly spaced proximally; rare or absent distally. Origin of M not preserved. MA dichotomously branched relatively close to wing margin. MP pectinately branched, with four branches, two deeply forked; one preserved crossvein evident between them. CuA long, pectinately branched with 11 long branches, at least one deeply forked; four-six crossveins between proximal branches forming gradate series. CuP incompletely preserved, short, apparently deeply forked. Anal veins not preserved. Crossveins between RP 1 and MA, MA and MP, MP and CuA rather scarce, irregularly spaced. Type material. Holotype No. 15453 (part; formerly No. 10336), No. 15454 (counterpart), deposited in TM. A poorly-preserved specimen with two incomplete hind wings. Type locality and horizon. Solnhofen, Germany; Late Jurassic, early Tithonian (Solnhofen Formation). Discussion. Handlirsch (1906–1908) thought that these are forewings, apparently following Weyenbergh (1869 a) and Haase (1890)., Published as part of Shi, Chaofan, Makarkin, Vladimir N., Yang, Qiang, Archibald, S. Bruce & Ren, Dong, 2013, New species of Nymphites Haase (Neuroptera: Nymphidae) from the Middle Jurassic of China, with a redescription of the type species of the genus, pp. 393-410 in Zootaxa 3700 (3) on pages 395-397, DOI: 10.11646/zootaxa.3700.3.4, http://zenodo.org/record/248765

Nymphites bimaculatus Shi, Makarkin et Ren, sp. nov. (Figs 3–8) Diagnosis. May be distinguished from Nymphites priscus by crossveins present between all branches of CuA, MP; from Nymphites sp. A by a more obtuse angle between Cu and its branches. Description. Holotype CNU-NEU-NN 2011116 PC. Thorax, abdomen fragmentarily preserved; details not visible. Metafemora relatively stout. Metatibiae long, slender with remnants of terminal spurs (Fig. 4 A); covered with dense, relatively short hairs. Tarsus covered with fine, short hairs; four proximal tarsomeres distally with row of strong dark bristles. Claws moderately large; not strongly curved; basally not strongly dilated. Forewing: left wing 37.5 mm long as preserved (estimated complete length 38 mm), 11 mm wide; right wing 33 mm as preserved, 12.2 mm wide. One trichosor between tips of each two veins along apical, outer margins of wings; three-four trichosors between tips of each two veins along hind margin in proximal half of wing (Fig. 5). Costal space narrow, strongly dilated after fusion of ScP, RA. Proximal subcostal veinlets mostly simple, few forked; all distal subcostal veinlets once or twice forked; no crossveins between them. ScP, RA stout for entire length. Subcostal space narrow, slightly dilated distally; no distinct crossveins detected (but see Remarks). Veinlets of ScP + RA dichotomously branched; no crossveins between them. RP smooth, not zigzagged; originating near wing base. Ten irregularly spaced crossveins between RA, RP before fusion of ScP + RA; not detected after fusion. RP with 14 (left wing), 13 (right wing) branches; all dichotomously branched (RP 1, RP 2 more deeply than others). Crossveins in radial space rather numerous, irregularly spaced in proximal two thirds; rare in distal portion, most form outer gradate series. M incurved; forked far proximad origin of RP 1. One preserved crossvein connecting RP, M before fork. MA, MP parallel. MA branched (probably dichotomously) relatively close to wing margin (incompletely preserved). MP zigzagged distally, pectinately branched; with four branches, two deeply forked (left wing), five shallowly forked branches (right wing); four crossveins between them. CuA long, pectinately branched with four long branches, two-three deeply forked; four crossveins between these. CuP long, parallel to posterior margin; pectinately forked: left wing, with six long branches, two proximal-most deeply forked; right wing, nine branches, four of them deeply forked; no crossveins between these. All preserved branches of CuA once or twice shallowly forked. CuP space twice wider than intracubital space. AA 3 + 4 long, pectinately branched, with two long branches, very shallowly forked (left wing); dichotomous, with anterior branch very shallowly forked, posterior branch deeply forked (right wing). AP 1 + 2 deeply forked; anterior branch forked, posterior at most shallowly forked (incompletely preserved). Crossveins between RP 1 and MA, MA and MP, MP and CuA, CuA and CuP rather numerous, irregularly spaced. Two crossveins between CuP, AA 3 + 4; one crossvein between AA 3 + 4, AP 1 + 2. Wing patterns consisted of two spots near hind wing margin, more or less rounded at MA, elongate at outer gradate series in radial space. Hind wing: left wing about 36 mm long as preserved (estimated complete length 37 mm), 10 mm wide; right wing 31.5 mm as preserved, 10.8 mm wide. Trichosors preserved between each vein pair terminating along margin, one medially, three-four basally (Fig. 5). Costal space narrower than in forewing. Subcostal veinlets mostly simple in proximal 2 / 3; once or twice shallowly forked distally. ScP, RA stout for entire length. Subcostal space narrow; no crossveins detected. Preserved veinlets of ScP + RA dichotomously branched; no crossveins connecting them detected. RP smooth, not zigzagged; originating near wing base. Eleven irregularly arranged crossveins between RA, RP before fusion of ScP + RA detected; not detected after fusion. RP with 11 branches; RP 1 to RP 3 deeply forked. Crossveins in radial space rather numerous, irregularly spaced in proximal proximally; rare or absent distally. Origin of M not preserved. MA dichotomously branched relatively close to wing margin. MP pectinate branched, with five branches, two-three deeply forked; three-four crossveins between them. CuA long, pectinately branched with 12 long branches, three-four deeply forked; 11–12 crossveins between them forming gradate series. All preserved branches of MP, CuA once or twice shallowly forked. CuP incompletely preserved, short, deeply forked. Anal veins not preserved. Crossveins between RP 1 and MA, MA and MP, MP and CuA rather numerous, irregularly spaced. Two-three crossveins between CuP and AA 3 + 4. Paratype CNU-NEU-NN 2011117 PC. Forewing 38.4 mm long, 11.5 mm wide as preserved (estimated complete length about 40–41 mm, width about 12 mm). Trichosors preserved in distal part of anterior wing margin. Costal space relatively narrow; strongly dilated after fusion of ScP, RA. Subcostal veinlets not preserved proximally; mainly forked medially; all once or twice shallowly forked distally; no crossveins connecting them. ScP, RA stout for entire length. Subcostal space moderately broad; slightly dilated distally; no crossveins detected. ScP + RA entering margin well after wing apex; its veinlets dichotomously branched; no crossveins connecting these. RP smooth not zigzagged; originating near wing base. Ten crossveins between RA, RP before fusion of ScP + RA widely spaced; not detected after fusion. RP with 11 branches; all dichotomously branched (RP 1, RP 3, RP 4 more deeply than others). Crossveins in radial space rather numerous, irregularly spaced in proximal two thirds; rare in distal portion, most form outer gradate series. M slightly incurved; forked far proximad origin of RP 1. One preserved crossvein connecting RP, M before fork. MA, MP parallel. MA branched (probably dichotomously) relatively close to wing margin. MP pectinately branched, with four branches; three crossveins between them. CuA long, pectinately branched with five long branches; four crossveins between them. CuP long, parallel to posterior margin; pectinate branched with six long branches; no crossveins between these. Branches of CuA, CuP incompletely preserved; some rather deeply forked. CuP space more than twice wider than intracubital space. Several irregularly spaced crossveins between CuA, CuP. AA 3 + 4 long, pectinately branched, with three branches; preserved branches shallowly forked; proximal branch deeply forked. AA 3 + 4, CuP closely approach basally. AP 1 + 2 deeply forked. Crossveins between RP 1 and MA, MA and MP, MP and CuA, CuA and CuP rather numerous, irregularly spaced. One crossvein between CuP and AA 3 + 4, AA 3 + 4 and AP 1 + 2, AP 1 + 2 and AP 3 + 4. Wing color pattern: two spots near hind wing margin, at MA, at outer gradate series in radial space. Hind wing about 37 mm long, 10.4 mm wide. Trichosors preserved in distal part of anterior wing margin. Costal space narrower than in forewing. Subcostal veinlets simple proximally, medially; once or twice shallowly forked distally. ScP, RA stout for entire length. Subcostal space moderately broad; slightly dilated distally; no crossveins detected. ScP + RA entering margin well after wing apex; its veinlets dichotomously branched; no crossveins connecting these. RP smooth, not zigzagged; originating near wing base. Nine crossveins between RA, RP before fusion of ScP + RA detected, widely spaced; not detected after fusion. RP with 12 branches; RP 1 to RP 4 deeply forked. Crossveins in radial space rather numerous, irregularly spaced in proximal proximally; rare distally, most of them form outer gradate series. Origin of M not preserved. MA branched relatively close to wing margin. MP pectinately branched, with four branches, two deeply forked; two crossveins between these. CuA long, pectinately branched, with eight long branches; all but one deeply forked; seven crossveins between them forming gradate series. CuP incompletely preserved, short, probably deeply forked. Anal veins not preserved. Crossveins between RP 1 and MA, MA and MP, MP and CuA rather numerous, irregularly spaced. One crossvein between CuP and AA 3 + 4, AA 3 + 4 and AP 1 + 2, AP 1 + 2 and AP 3 + 4. Material. Holotype CNU-NEU-NN 2011116 PC (part, counterpart), four well preserved wings and part of a poorly preserved thorax and abdomen. Paratype CNU-NEU-NN 2011117 PC (part, counterpart), a forewing and hind wing, overlapping. Both specimens are deposited in CNUB. Etymology. The specific epithet is formed from the Latin bi, two, and maculatus, spotted, in reference to the presence of two small spots on the forewing. Type locality and horizon. Daohugou Village, Wuhua Township, Ningcheng County, Inner Mongolia, China. Middle Jurassic, Bathonian/Callovian; Jiulongshan Formation. Remarks. The new species is assigned to Nymphites based on the strong similarity of its hind wing venation to that of the type species N. priscus, e.g., irregularly arranged crossveins in the radial space; the four pectinate branches of MP; the presence of a series of gradate crossveins between branches of CuA; short CuP. Nevertheless, this assignment is preliminary because of the incompleteness of the holotype. The holotype and paratypes of Nymphites bimaculatus sp. nov. have similar venation in general, and identical forewing patterning (two small spots). Their venation differs in details, but these differences do not extend beyond those of the holotype left and right wings. Although subcostal crossveins in this species are not confidently detected, we assume that this is likely an artefact of preservation, and that a few fine crossveins (not distinctly visible) and features of thyridiate crossveins (ScP with small scarce projections posteriorly) are probably present. We explain the clear difference in width noted between the right (wider) and left (narrower) wings as a result of post-mortem plastic distortion, extension of the matrix approximately along the plane of the length of the left wings. Therefore, we estimate that the actual forewing length of the holotype is probably 34–35 mm (not 37 mm)., Published as part of Shi, Chaofan, Makarkin, Vladimir N., Yang, Qiang, Archibald, S. Bruce & Ren, Dong, 2013, New species of Nymphites Haase (Neuroptera: Nymphidae) from the Middle Jurassic of China, with a redescription of the type species of the genus, pp. 393-410 in Zootaxa 3700 (3) on pages 398-403, DOI: 10.11646/zootaxa.3700.3.4, http://zenodo.org/record/248765

Nymphidae Rambur, 1842, Published as part of Shi, Chaofan, Makarkin, Vladimir N., Yang, Qiang, Archibald, S. Bruce & Ren, Dong, 2013, New species of Nymphites Haase (Neuroptera: Nymphidae) from the Middle Jurassic of China, with a redescription of the type species of the genus, pp. 393-410 in Zootaxa 3700 (3) on page 395, DOI: 10.11646/zootaxa.3700.3.4, http://zenodo.org/record/248765

Nymphites sp. A (Figs 4 B, 9, 10) Description. Thorax, abdomen fragmentary preserved; details not visible. Meso-, metafemora long, rather swollen. Mesotibiae relatively stout, short. Metatibiae long, slender with short terminal spurs (Fig. 4 B). All preserved segments covered with dense, relatively short hairs. Hind wing 25.8 mm long, 10.2 mm wide as preserved. Trichosors not preserved. Costal space rather narrow. Subcostal veinlets mostly simple in proximal 2 / 3; once or twice shallowly forked distally. ScP, RA stout for entire length. Subcostal space moderately broad; no crossveins detected. RP smooth, not zigzagged; originating near wing base. Eleven irregularly arranged crossveins between RA, RP before fusion of ScP + RA; not detected after fusion. RP with eight preserved branches (estimated 11–12 complete). Crossveins in radial space rather rare, irregularly spaced in proximal proximally. MA, MP fragmentarily preserved, branching not preserved. Intermedial crossveins long, somewhat oblique. CuA long, pectinate branched with nine preserved long branches, some deeply forked; branches inclined to CuA at rather acute angle; all preserved branches connect by one crossveins forming gradate series. CuP fragmentarily preserved, short. Anal veins not preserved. Material. CNU-NEU-NN 2011118 PC (part, counterpart), deposited in CNUB. Fragmentarily preserved body with two incomplete hind wings. Remarks. This species is clearly not conspecific with Nymphites bimaculatus sp. nov. It differs from the latter by the more acute angle between Cu and its branches, more scarce crossveins, and probably wider hind wing. However, as this specimen is very incompletely preserved, we refrain from naming the species pending more complete specimens.

Genus Nymphites Haase, 1890 Nymphites Haase, 1890: 23; Handlirsch 1906–1908 [1906]: 608 [Nymphitidae]; Jepson et al. 2012: 44 [Nymphidae]. Type species. Hemerobius priscus Weyenbergh, 1869 a, by original designation. Diagnosis. Forewing elongate, broad proximally [strongly narrowed proximally in Liminympha Ren et Engel, 2007]; all distal veinlets of ScP once or twice forked [all veinlets of ScP simple in Mesonymphes Carpenter, 1929]; MP with four-five pectinate branches, each connected by one crossvein [crossveins absent between branches of MP in Liminympha]; CuP space broad, more than twice as wide as intracubital space [narrow, nearly as wide as intracubital space in Liminympha]. Hind wing MP with four-five pectinate branches [eight branches in Sialium Westwood, 1854]; at least proximal branches of CuA connected by crossveins [not connected in Mesonymphes]; CuP short, not pectinately branched. Legs (at least hind) with tibial spurs. Composition. Two species from the Jurassic of Eurasia: Nymphites bimaculatus sp. nov. from the Middle Jurassic of China, and Nymphites priscus (Weyenbergh, 1869) from the Late Jurassic of Germany., Published as part of Shi, Chaofan, Makarkin, Vladimir N., Yang, Qiang, Archibald, S. Bruce & Ren, Dong, 2013, New species of Nymphites Haase (Neuroptera: Nymphidae) from the Middle Jurassic of China, with a redescription of the type species of the genus, pp. 393-410 in Zootaxa 3700 (3) on page 395, DOI: 10.11646/zootaxa.3700.3.4, http://zenodo.org/record/248765

Shi, Chaofan, Makarkin, Vladimir N., Yang, Qiang, Archibald, S. Bruce, Ren, Dong (2013): New species of Nymphites Haase (Neuroptera: Nymphidae) from the Middle Jurassic of China, with a redescription of the type species of the genus. Zootaxa 3700 (3): 393-410, DOI: http://dx.doi.org/10.11646/zootaxa.3700.3.4

A subgroup of leukemogenic mixed-lineage leukemia (MLL) fusion proteins (MFPs) including MLL-AF9 activates the Mecom locus and exhibits extremely poor clinical prognosis. Mecom encodes EVI1 and MDS1-EVI1 (ME) proteins via alternative transcription start sites; these differ by the presence of a PRDI-BF1-RIZ1 (PR) domain with histone methyltransferase activity in the ME isoform. Using an ME-deficient mouse, we show that ME is required for MLL-AF9-induced transformation both in vitro and in vivo. And, although Nup98-HOXA9, MEIS1-HOXA9, and E2A-Hlf could transform ME-deficient cells, both MLL-AF9 and MLL-ENL were ineffective, indicating that the ME requirement is specific to MLL fusion leukemia. Further, we show that the PR domain is essential for MFP-induced transformation. These studies clearly indicate an essential role of PR-domain protein ME in MFP leukemia, suggesting that ME may be a novel target for therapeutic intervention for this group of leukemias.

EVI1 is a zinc finger oncoprotein that binds via fingers 1-7 to the sequence GACAAGATAA. The target genes on which EVI1 acts are unknown. This binding motif overlaps with that for the GATA transcription factors, (T/A)GATA(A/G), and GATA-1 can bind to and activate transcription via a GACAAGATAA motif. The possibility has been raised that, when overexpressed in leukemogenesis, EVI1 may function by interfering with the differentiation-promoting action of GATA factors. To explore this, we have assessed the affinity of EVI1 for the GATA binding sites derived from erythroid-specific GATA-1 target genes, and found only low affinity interactions. We examined the contacts between EVI1 and DNA by methylation interference studies, which revealed extensive contacts between EVI1 and its binding site. The importance of the contacts for high affinity binding was shown by in vitro quantitative gel shift studies and in vivo cotransfection studies. To examine what types of sequences from mouse genomic DNA bind to EVI1, we isolated and sequenced five EVI1-binding fragments, and each showed the GACAAGATA site. The data presented contribute to our knowledge of the binding specificity of EVI1, and yield a clearer picture of what sequences can, and cannot, act as targets for EVI1 action.

The effects of climatic factors and vegetation type on evapotranspiration (E) and water use efficiency (WUE) were analyzed using tower-based eddy-covariance (EC) data of eleven African sites (22 site years) located across a continental-scale transect. The seasonal pattern of E was closely linked to growing-season length and rainfall distribution. Although annual precipitation (P) was highly variable among sites (290 to 1650 mm), minimum annual E was not less than 250 mm and reached a maximum of 900 mm where annual P exceeded 1200 mm. Site-specific interannual variability in E could be explained by either changes in total P or variations in solar irradiance. At some sites, a highly positive linear correlation was found between monthly sums of E and net radiation (Rn), whereas a hysteretic relationship at other sites indicated that E lagged behind the typical seasonal progression of Rn. Results of a cross-correlation analysis between daily (24-h) E and Rn revealed that site-specific lag times were between 0 days and up to a few weeks depending on the lag of vapor pressure deficit (D) behind Rn and vegetation type. Physiological parameters (e.g. mean dry-foliage Priestley-Taylor alpha) implied that stomatal limitation to transpiration prevailed. During the rainy season, a strong linear correlation between monthly mean values of gross primary production (GPP) and E resulted in water use efficiency being constant with lower values at grass-dominated sites (~2 to ~3.5 g C kg-1 H2O) than at natural woodland sites and plantations (~4.5 to ~6 g C kg-1 H2O).

Abnormal expression of the transcription factor EVI1 through chromosome 3q26 rearrangements has been implicated in the development of one of the most therapeutically challenging high-risk subtypes of acute myeloid leukemia (AML). Here we integrated genomic and metabolic screening of hematopoietic stem cells to reveal that EVI1 overexpression altered cellular metabolism. A pooled shRNA screen targeting metabolic enzymes identified the ATP-buffering, mitochondrial creatine kinase CKMT1 as a druggable dependency in EVI1-positive AML. Of 18 screened AML cell lines harboring various genetic alterations, only the four EVI1-expressing lines exhibited markedly elevated CKMT1 protein expression and activity. Treatment of this cell line panel with either CKMT1-targeting shRNAs or cyclocreatine, an analog of the CKMT1 substrate creatine and inhibitor of the creatine biosynthesis pathway, showed that elevated CKMT1 protein expression correlated with sensitivity to CKMT1 pathway inhibition. Consistent with these data, flow cytometry analysis of a panel of 68 unselected primary AML patient specimens revealed that the four leukemias with the highest levels of EVI1 expression also had elevated CKMT1 protein levels and enhanced sensitivity to cyclocreatine treatment. We next established that enforced EVI1 expression increased CKMT1 protein and mRNA levels and that three independent shRNA molecules targeting EVI1 drastically reduced CKMT1 expression in two EVI1-positive AML cell lines. A luciferase-based reporter system established that RUNX1 represses CKMT1 expression through direct binding to its promoter. ChIP-qPCR approaches were then applied to dissect the sequential events involved in EVI1-induced CKMT1 upregulation and the possible role of RUNX1 as an intermediate. In both primary AML samples and cell lines, we determined that EVI1 represses RUNX1 expression by directly binding to its promoter. This, in turn, eliminates repressive RUNX1 binding at the CKMT1 promoter and thereby promotes CKMT1 expression. Based on these data, we explored the relationship between EVI1 and RUNX1 expression with CKMT1 mRNA levels in two AML transcriptional datasets (GSE14468 and GSE10358). We divided these cohorts into four subgroups with high versus low expression of EVI1 and RUNX1. Consistent with our mechanistic analysis, primary AML samples within the EVI1high/RUNX1low subgroup were significantly more likely to express high levels of CKMT1 than AML samples in the other three subgroups. CKMT1 promotes the metabolism of arginine to creatinine. To determine the effect of CKMT1 suppression on this pathway, we measured the metabolic flux of stable-isotope labeled L-arginine 13C6 through creatine synthesis using mass spectrometry. CKMT1-directed shRNAs or cyclocreatine selectively decreased intracellular phospho-creatine and blocked production of ATP by mitochondria. Salvage of the creatine pathway by exogenous phospho-creatine restored normal mitochondrial function, prevented the loss of viability of human EVI1-positive AML cells induced by cyclocreatine or CKMT1-directed shRNAs, and maintained the serial replating activity of Evi1-transformed bone marrow cells. Primary human EVI1-positive AML is frequently associated with somatic NRAS mutations. Thus, to investigate whether EVI1 over-expression sensitizes primary AMLs to CKMT1 inhibition in vivo, we transplanted primary NrasG12D mutant AMLs with and without elevated Evi1 expression into congenic recipient mice. In this system, Ckmt1 knockdown did not significantly alter the outgrowth of control Nras mutant AML cells compared to a shControl (63% versus 71%). By contrast, NrasG12D AML cells characterized by elevated Evi1 expression were profoundly depleted by Ckmt1 suppression to 2% versus 58% in shControl recipients. Consistent with these results, pharmacologic or genetic inhibition of the CKMT1-dependent pathway blocked disease progression and prolonged the survival of mice injected with human EVI1-positive cells but not with EVI1-negative cells, without noticeable cytotoxic effect on normal murine cells. In conclusion, we have integrated "omic" approaches to identify CKMT1 as a druggable liability in EVI-positive AML. This study supports a potential therapeutic avenue for targeting the creatine kinase pathway in EVI1-positive AML, which remains one of the worst outcome subtypes of AML. Disclosures DeAngelo: Incyte: Consultancy; Novartis: Consultancy; Celgene: Consultancy; Amgen: Consultancy; Baxter: Consultancy; Pfizer: Consultancy; Ariad: Consultancy. Stone:Pfizer: Consultancy; Agios: Consultancy; Jansen: Consultancy; Celator: Consultancy; Merck: Consultancy; Amgen: Consultancy; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Karyopharm: Consultancy; Novartis: Consultancy; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Xenetic Biosciences: Consultancy; Sunesis Pharmaceuticals: Consultancy; Seattle Genetics: Consultancy; Roche: Consultancy; Juno Therapeutics: Consultancy; ONO: Consultancy.

The ecotropic virus integration site 1 (EVI1) transcription factor is associated with human myeloid malignancy of poor prognosis and is overexpressed in 8-10% of adult AML and strikingly up to 27% of pediatric MLL-rearranged leukemias. For the first time, we report comprehensive genomewide EVI1 binding and whole transcriptome gene deregulation in leukemic cells using a combination of ChIP-Seq and RNA-Seq expression profiling. We found disruption of terminal myeloid differentiation and cell cycle regulation to be prominent in EVI-induced leukemogenesis. Specifically, we identified EVI1 directly binds to and downregulates the master myeloid differentiation gene Cebpe and several of its downstream gene targets critical for terminal myeloid differentiation. We also found EVI1 binds to and downregulates Serpinb2 as well as numerous genes involved in the Jak-Stat signaling pathway. Finally, we identified decreased expression of several ATP-dependent P2X purinoreceptors genes involved in apoptosis mechanisms. These findings provide a foundation for future study of potential therapeutic gene targets for EVI1-induced leukemia.

The zinc finger protein EVI1 is causally associated with acute myeloid leukemogenesis, and inhibition of its function with a small molecule therapeutic may provide effective therapy for EVI1-expressing leukemias. In this paper we describe the development of a pyrrole–imidazole polyamide to specifically block EVI1 binding to DNA. We first identify essential domains for leukemogenesis through structure–function studies on both EVI1 and the t(3;21)(q26;q22)-derived RUNX1-MDS1-EVI1 (RME) protein, which revealed that DNA binding to the cognate motif GACAAGATA via the first of two zinc finger domains (ZF1, encompassing fingers 1–7) is essential transforming activity. To inhibit DNA binding via ZF1, we synthesized a pyrrole–imidazole polyamide 1, designed to bind to a subsite within the GACAAGATA motif and thereby block EVI1 binding. DNase I footprinting and electromobility shift assays revealed a specific and high affinity interaction between polyamide 1 and the GACAAGATA motif. In an in vivo CAT reporter assay using NIH-3T3-derived cell line with a chromosome-embedded tet-inducible EVI1-VP16 as well as an EVI1-responsive reporter, polyamide 1 completely blocked EVI1-responsive reporter activity. Growth of a leukemic cell line bearing overexpressed EVI1 was also inhibited by treatment with polyamide 1, while a control cell line lacking EVI1 was not. Finally, colony formation by RME was attenuated by polyamide 1 in a serial replating assay. These studies provide evidence that a cell permeable small molecule may effectively block the activity of a leukemogenic transcription factor and provide a valuable tool to dissect critical functions of EVI1 in leukemogenesis.

Recent studies have indicated a role for a MECOM allele in susceptibility to osteoporotic fractures in humans. We have generated a mutation in Mecom in mouse (termed ME(m1)) via lacZ knock-in into the upstream transcription start site for the gene, resulting in disruption of Mds1 and Mds1-Evi1 transcripts, but not of Evi1 transcripts. We demonstrate that ME(m1/m1) mice have severe kyphoscoliosis that is reminiscent of human congenital or primary kyphoscoliosis. ME(m1/m1) mice appear normal at birth, but by 2weeks, they exhibit a slight lumbar lordosis and narrowed intervertebral space. This progresses to severe lordosis with disc collapse and synostosis, together with kyphoscoliosis. Bone formation and strength testing show that ME(m1/m1) mice have normal bone formation and composition but are osteopenic. While endochondral bone development is normal, it is markedly dysplastic in its organization. Electron micrographs of the 1week postnatal intervertebral discs reveals marked disarray of collagen fibers, consistent with an inherent weakness in the non-osseous connective tissue associated with the spine. These findings indicate that lack of ME leads to a complex defect in both osseous and non-osseous musculoskeletal tissues, including a marked vertebral osteopenia, degeneration of the IVD, and disarray of connective tissues, which is likely due to an inherent inability to establish and/or maintain components of these tissues.

A component of a heterodimeric cytochrome b, designated gp91-phox, is required for the microbicidal activity of phagocytic cells and is expressed exclusively in differentiated myelomonocytic cells (granulocytes; monocyte/macrophages). In an attempt to identify cis-elements responsible for this restricted pattern of expression, we produced transgenic mice carrying reporter genes linked to the human gp91-phox promoter. Immunohistochemical and RNA analyses indicate that 450 base pairs of the proximal gp91-phox promoter is sufficient to target reporter expression to a subset of monocyte/macrophages. Mice expressing simian virus 40 large tumor antigen under control of the gp91-phox promoter develop monocyte/macrophage-derived malignancies with complete penetrance at 6-12 mo of age and provide an animal model of true histiocytic lymphoma. As these transgenes are inactive in most phagocytic cells that express the endogenous gp91-phox-encoding gene, we infer that additional genomic regulatory elements are necessary for appropriate targeting to the full complement of phagocytes in vivo.

Evi-1 was originally identified as a common site of viral integration in murine myeloid tumors. Evi-1 encodes a 120-kDa polypeptide containing 10 zinc finger motifs located in two domains 380 amino acids apart and an acidic domain located carboxy terminal to the second set of zinc fingers. These features suggest that Evi-1 is a site-specific DNA-binding protein involved in the regulation of RNA transcription. We have purified Evi-1 protein from E. coli and have employed a gel shift-polymerase chain reaction method using random oligonucleotides to identify a high-affinity binding site for Evi-1. The consensus sequence for this binding site is TGACAAGATAA. Evi-1 protein specifically protects this motif from DNase I digestion. By searching the nucleotide sequence data bases, we have found this binding site both in sequences 5' to genes in putative or known regulatory regions and within intron sequences.

This review will begin with a detail of the revision of the WHO classification, and pathological definitions of Burkitt lymphoma. Over the past several years, molecular understanding of Burkitt lymphoma has improved significantly. Using gene expression profiling, a genomic “signature” of Burkitt lymphoma may be identified, that has fidelity beyond c-myc expression, and the presence of the classical t(8;14). Then, evaluation and therapy of the adult patient with Burkitt lymphoma will be reviewed. Relatively few data exist on optimal therapy of the adult patient with Burkitt lymphoma. Principles of therapy should include high doses of alkylating agents, frequent administration of chemotherapy, and attention to central nervous system (CNS) prophylaxis with high doses of systemic chemotherapy, intrathecal therapy, or both. The outcome of adult patients with Burkitt lymphoma, particularly those over 40 years of age, is inferior to the outcome of younger patients, but may be improving over the past few years. Results from an international collaborative effort, which are helpful in evaluating results of Burkitt lymphoma therapy in adults, will be presented. HIV-associated Burkitt lymphoma, and elderly patients with Burkitt lymphoma, comprise special clinical situations that will be also covered in this review.

Purpose Midline sternotomy remains the preferred technique for access in cardiac surgery. Application of steel wires has been the preferred method of closure. Because of associated complications, such as superficial and deep infections, as well as bony nonunion complications, an alternative technique is being proposed. The purpose of this study is to evaluate results of a new device for sternal closure. Description The Sternal Talon (KLS Martin Group, Jacksonville, FL), a lightweight titanium closure device is designed to encircle the sternum, thus yielding a stable closure by effectively distributing the strength of closure over the entire length of the sternotomy. After multiple strength tests demonstrated its superiority over wires, and cadaver tests confirmed its ease of placement, the Food and Drug Administration recently approved the device for its unrestricted use. Eight institutions were chosen to perform initial placements. Patient selection was limited to patients at high risk for sternotomy complications. Evaluation In 42 patients who underwent placement of the Sternal Talon (KLS Martin Group) after sternotomy, no wound infections or dehiscence, nonunions, or returns to the operating room were observed. Three postoperative deaths were reported, none of which were device related. The device is magnetic resonance imaging compatible and there are no reported problems with computed tomographic scatter or chest roentgenogram visualization. Conclusions These initial cases prove the safety and efficacy of the Sternal Talon device for sternum closure in high-risk patients and may be regarded as an alternative to conventional wire closure. Future prospective studies are warranted to prove the superiority of the device in terms of long-term stability and sternum union rates, as well as decreased infection rates specifically in the high-risk patient population undergoing sternotomy.

Chromosomal abnormalities, such as translocation, mutation or deletion, are central to the pathogenesis of human cancers. Recently, several transcription factors have been isolated as genes responsible for leukemia from the region surrounding chromosomal breakpoints, which are implicated in the regulation of normal hematopoiesis. Among on them, ecotropic viral integration site-1 (Evi1) is a transcription factor activated by retroviral integration in murine leukemias and chromosomal rearrangements in human leukemias. Evi1 is a zinc finger transcription factor and contains two separated DNA-binding domains. It was reported that Evi1−/− embryos die at approximately E10.5, exhibiting widespread hypocellularity and hemorrhaging. However, the role in normal hematopoiesis or authentic target genes of Evi1 has not been elucidated. Here, we show that Evi1 is predominantly expressed in hematopoietic stem cells (HSCs) in embryos and adult bone marrows, and Evi1−/− embryos are markedly decreased in numbers of HSC. One embryo-equivalent cells from E9.5 P-Sp of Evi1+/+, Evi1+/− and Evi1−/− embryos (Ly5.2) were transplanted into a busulfan-conditioned newborn recipient (Ly5.1). At 2 months posttransplant, donor-derived Ly5.2(+) cells could be detected in the peripheral blood of the recipients that received P-Sp cells from the Evi1+/+ and Evi1+/− but not from the Evi1−/− embryos. Thus, Evi1 is critical for the generation of HSCs in the P-Sp. Both Evi1−/− embryos and yolk sac showed marked retardation in the organization of the vascular system, particularly in vascular remodeling, compared with controls. Using an in vitro P-Sp culture analysis, we found normal in vitro differentiation of endothelial cells in Evi1−/− P-Sp cultures but defects in their in vitro network formation, which is normally promoted by Ang-1 secreted from developing HSCs in P-Sp cultures. HSCs from adult bone marrow or HSCs from E9.5 wild type embryos rescued defective angiogenesis in Evi1−/− embryos. The fine vascular network coincided with the region where HSCs formed a colony. Their round morphology confirmed that exogenous adult HSCs did not differentiate into elongated endothelial cells. We showed that recombinant Ang-1 alone restored the defective angiogenesis in Evi1−/− embryos to a wild type level. It is suggested that the defect in hematopoietic cells induced defective angiogenesis in Evi1−/− embryos mediated by Ang-1. Notably, mRNA expression of GATA-2, which is essential for proliferation of definitive HSCs, was profoundly reduced in Evi1−/− embryos. Analysis of the GATA-2 promotor revealed that Evi1 directly binds to the 5′ upstream region of the GATA−2 exon and positively regulates its promoter activity in vitro and in vivo. Restoration of GATA-2 expression dramatically rescued the defective expansion of Evi1−/− embryos HSCs in vitro. Our results reveal that GATA-2 is a critical in vivo target for Evi1 and indicate hierarchical regulation of the HSC pool by transcriptional regulators.