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To the Editor: In a recent issue of Blood, Pane et al[1][1] reported on five cases of mild Phi+ myeloproliferative disorders characterized by a c3/a2 junction in the BCR/ABL hybrid gene transcript and suggested to use the term “neutrophilic chronic myeloid leukemia (N-CML)” to distinguish a “

Protein engineering has made it possible to combine the binding sites of murine antibodies with human antibody regions. Antibodies constructed in this way have important advantages for therapy.

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The usefulness of isozyme and protein markers in identifying tilapia species and their hybrids is demonstrated Genetic characterization of Oreochromis niloticus populations from commercial farms, experimental stations, and government hatcheries in Luzon, Visayas and Mindanao in the Philippines indicates well-established introgression with O mossambicus Genetic differentiation of the O niloticus stocks, measured by Nei's genetic distance, was highly correlated with O mossambicus gene content The implications of these results for tilapia genetic improvement are discussed.

Cord blood samples of 113 Japanese babies from Kurashiki, Japan, were analyzed for the presence of possible modifications in gamma-globin gene arrangements and for changes in the relative quantities of the gamma chains G gamma, A gamma, and its variant A gamma T. As many as 9 babies had the gamma-globin gene triplication, one a gamma-globin gene quadruplication, and two a gamma-globin gene deletion. The triplication and quadruplication involved cross-overs within the G gamma- and A gamma-globin genes resulting in a hybrid gene with the 5'-segment being derived from the A gamma and the 3'-segment from the G gamma-globin gene (-AG gamma-). The G gamma value of the 9 babies with the -G gamma-AG gamma-A gamma-globin gene arrangement averaged approximately 80% and that of the baby with -G gamma-AG gamma-AG gamma-A gamma- was 78%. Four of the 9 babies with the triplication and the baby with the quadruplication had an A gamma T mutation in trans; in these babies, the A gamma chain level (from the A gamma-globin gene in 3'-position in the tri- and quadruplication arrangements) was decreased to about one-third and one-sixth of the level observed in the simple A gamma T heterozygote. This observation supports the suggestion that transcription beginning at the 5'-gamma-globin gene interferes with that of the 3'-gamma-globin gene.

Two species of tilapia, Oreochromis mossambicus (Peters) and O. niloticus (L.), from five reservoirs in southern Sri Lanka as well as fish from a fish hatchery and laboratory stock were studied by starch gel electrophoresis at 18 enzyme loci, to determine the degree of mixing of the two species. Two reservoirs showed little or no mixing between O. mossambicus and O. niloticus, while two other reservoirs as well as the hatchery stock showed a high degree of mixing of the two species. The laboratory stock of O. niloticus also showed introgression of O. mossambicus alleles. The similarities of the local stocks to the original African as well as other Asian stocks is discussed.

The ability to measure gene expression with light is revolutionizing biotechnology. It is now possible to visualize transcription or translation and localize it to particular domains, cells or organelles of any organism. Moreover, this can be done repeatedly, noninvasively, and with unsurpassed sensitivity.

Since HIV tat function is essential for the HIV infectious cycle, it represents an important possible target of therapeutic intervention for HIV infection. Stable human cell lines were derived that express high levels of beta-galactosidase under the combined control of the transacting HIV-1 tat gene product and the cis-acting HIV-1 LTR. The tat gene product induces LTR-linked gene expression approximately 1000-fold in this system. The high level of expression of beta-galactosidase under HIV tat and LTR control in stable cell lines allows rapid spectrophotometric quantitation of beta-galactosidase enzymatic activity from fewer than 5000 cells seeded in a microtiter plate well. Such cell lines provide a virus-free system for the high-capacity screening of compounds for the ability to interfere with HIV tat-mediated transactivation of gene expression.

Hybrid proteins consisting of N-terminal fragments of increasing length of a Bacillus thuringiensis insecticidal protein (Bt2) fused to neomycin phosphotransferase II (NPTII) were produced in Escherichia coli. The minimum fragment required for insect toxicity is comprised of the first 607 amino acids of Bt2. Fusion proteins not containing this minimum fragment were non-toxic. The NPTII activity of the different non-toxic hybrid proteins varied considerably but was not correlated with the length of the Bt2 fragment. Fusion proteins including the minimum toxic fragment of Bt2 exhibited insecticidal and NPTII activity comparable to that of the individual proteins. This was largely independent of the fusion point within Bt2. Our data suggest that the conformation of the Bt2 polypeptide exerts an important influence on the enzymatic activity of the fused NPTII protein. The combination of insecticidal activity and a dominant selectable trait into one protein offers important advantages for the generation of insect resistant transgenic plants.

By the phosphitetriester approach we synthesized the fragments for constructing a synthetically idealized promoter (SIP). The total construction consists of the promoter, operator and Shine-Dalgarno (SD) region. This control region was fused to the gene for γ-interferon. We chemically synthesized a control region in order to optimally express our synthetic genes considering the following criteria. For easy modifications, e.g. for structure function studies we designed each functional domain as a cassette which can conveniently be dissected by the appropriate restriction enzymes. The regulation unit consists of a promoter1,2, an operator3 and a Shine-Dalgarno-sequence4 (FIG. 1).

A psbA gene fragment has been cloned into an expression vector in frame with the 3′ end of the beta-galactosidase gene. Expression of this construct in E. coli results in an overproduction of a hybrid protein consisting of part of the herbicide binding protein and beta-galactosidase. An antiserum raised against this fusion protein specifically detects a 34 kDa polypeptide within the complex mixture of spinach thylakoid membrane proteins.

Median duration of the chronic phase of chronic myelogenous leukemia (CML) is 3 years; less than 20% of patients have a chronic phase greater than 7 years. It is unknown whether the length of chronic phase is stochastic or is predetermined for each patient. Since molecular abnormalities of bcr and c-abl occur in CML, we sought to determine whether there were differences in bcr and c-abl translocation or transcription in individuals with long v short chronic phase. These studies were performed in six patients with CML in whom chronic phase was 7+ to 26 years and 20 patients in whom chronic phase was less than 7 years. All patients had translocation of c-abl to within bcr. The distribution of breakpoints in bcr were similar in both groups. Transcription of the chimeric bcr/c-abl mRNA was comparable. These data suggest that changes in bcr or c-abl alone do not determine the duration of chronic phase in CML; other factors are likely involved.

Revue des donnees biochimiques sur la localisation des proteines chez les cellules eucaryotes et chez Escherichia coli. Revue des techniques genetiques utilisees pour l'etude de la localisation des proteines chez les bacteries

A method is described for achieving the efficient production of proteins of biological interest by the establishment of hybridomas from lymphocytes of transgenic animals carrying a fusion gene having promoter-regulatory sequences functional in lymphocytes fused to the coding sequences of the protein to be produced. While possible improvements in the method are discussed, it is anticipated that the method will ultimately be applicable to the production of any protein of interest.

EXPERIMENTA L A PPROA CHES TO MAM MA LIA N GENE REGULA TIO N ....... . 274 HYBRID GENE MET HODS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 274 Cloned Gene...... . . . . . ......... ..... . .. 275 Expression System ...... . . . . . ......... .. . . .. 275 Hybrid Gene Construction........ . . . . . ...... 275 EXPERIMENTAL S ySTEMS . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . 277 Erythropoietic Gene Control... . .. . . . . ......... . . . 277 Myogenesis ..... . . ... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 281 Liver Specific Genes . . . . . ... . . ........ . ..... 284 Pancreatic Genes.. . .... . . . . .... . .... . . . . . . . . . . .... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 286 Miscellaneous...... . . . ..... 288 W HAT HAVE WE LEA RNED ? C ONCL USI ONS AND DIRECTI ONS . . . . . . . . ........ . . . 288 Cis-Acting Regulatory Sequences ....... . . . 288 Trans-Acting Factors...... . . . . . . . . . ........ ... . . . . .... ... . . ... . . . . . . ..... . ....... 289

Article synthese sur la structure moleculaire des antigenes d'histocompatibilite de classe I murins et humains. Des molecules ayant des mutations et substitutions de certains aminoacides ainsi que les produits de l'expression de genes hybrides ont ete etudies pour leur reconnaissance par les cellules T

The expression of the Escherichia coli uvrD gene was studied with a uvrD::Mud(Aprlac) insertion mutant. The results indicate that it is inducible by DNA damaging agents in a recA+ gene-dependent manner.

Article de synthese sur les tumeurs associees aux oncogenes transgeniques et utilisation des souris transgeniques en vue d'etudier les etapes de la tumorigenese

Description des differentes classes d'interferon, methodes de clonage des genes et de synthese chez les microorganismes; techniques de production d'analogues: structure et proprietes therapeutiques eventuelles

In yeast alpha cells, the product encoded by the MAT alpha 1 gene of the mating-type locus is required for transcription of at least two genes, STE3 and MF alpha 1. To learn whether the lack of the MAT alpha 1 product in a and a/alpha cells is sufficient to explain the failure to express STE3 and MF alpha 1 in these cells, we have provided MAT alpha 1 product via a hybrid gene that circumvents the normal regulation of the MAT alpha 1 gene. We find by RNA blot analysis that provision of MAT alpha 1 protein permits production of STE3 and MF alpha 1 mRNA in a cells but not in a/alpha cells. These data suggest the existence of an additional regulatory mechanism that prevents expression of alpha-specific genes in a/alpha cells, even when MAT alpha 1 product is present. This regulatory mechanism appears to control expression of STE3 and MF alpha 1 at the transcriptional level rather than at the posttranscriptional level, because we show that MF alpha 1 mRNA supplied from a constitutive promoter is translated and processed in a/alpha cells to yield functional alpha-factor pheromone. This result shows further that a/alpha cells possess all the machinery necessary for pheromone maturation, even though these cells do not normally carry out these reactions.

The copy number of R1 and of a repA - lacZ gene fusion was increased above normal by coupling to a plasmid which is present at a fivefold higher copy number at 30 °C. This carrier plasmid is deficient in replication at 42 °C, and it was thus possible after a temperature shift to analyze the response to the increased plasmid concentration of the R1 replication control system. Both the frequency of replication per plasmid molecule and the rate of repA expression per gene copy were reduced under these conditions, and the data strongly suggest that there is an inverse proportionality between the specific rate of plasmid replication viz repA expression and the copy number/gene dosage of the plasmid.

The p53 tumour suppressor induces cell cycle arrest and apoptosis in response to cellular stresses. p53 is inactivated by various cellular and viral factors. We set out to generate regulatable p53 derivatives that are highly inducible by synthetic ligands, escape inactivation and efficiently induce apoptosis. We have generated Ligand Inducible Chimeric Tumour Suppressors (LI-CTS), that are inactive unless provided with artificial ligands. They are resistant to inactivation, due to the replacement of domains that mediate p53 inhibition by heterologous sequences. LI-CTS are activated by micromolar concentrations of ligand in a variety of cell lines. Following ligand addition, they translocate to the nucleus, activate p53 inducible genes and induce apoptosis. We have established human head and neck squamous cell carcinoma lines that stably express LI-CTS, which are inducible. These lines form tumours in nude mice in the absence of ligand. Addition of ligand inhibits tumour formation, and moreover, regresses established tumours by apoptosis. Although regulatable p53 expression has been achieved previously, our study provides the first demonstration of regulatable in vivo regression of tumours in a p53 based approach. Regulated inhibition and regression of tumours with a ligand inducible chimeric tumour suppressor could provide a novel approach to p53 based gene therapy.

Resultats des experiences menees par l'equipe de Schell et Von Montagu a Ghent en collaboration avec le groupe de Cashmore a l'Universite Rockfeller consistant en la construction d'un gene chimere entre le gene nucleaire de la petite sous-unite de la RuBPC de pois et la sequence codante d'un gene bacterien de la chloramphenicol acetyltransferase. Apres incorporation dans un plasmide Ti fournissant les signaux de terminaison et de polyadenylation et infection de plants de tabac, l'expression du gene cat dans les chloroplastes matures est regulee par la lumiere

PhoE, one of the outer membrane proteins in E. coli K-12 cells, is involved in the uptake of small hydrophilic, especially negatively charged solutes. Its synthesis is induced upon phosphate limitation. Several models have been proposed for the mechanism of transport and incorporation of outer membrane proteins into the outer membrane. Escherichia coli cells containing a hybrid phoE/lacZ gene were studied to investigate the destination of the hybrid gene product, using an immunocytochemical labeling procedure and ultra-thin cryosectioning.

Essais de transformation directe de protoplastes de mesophylle de tabac par un plasmide d'E. coli ou est insere le gene de l'aminoglycoside phosphotransferase lie au promoteur et aux sequences de terminaison du gene VI du CaMV