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Here, we report features of a novel transcription factor zfh-5, which we isolated from the mouse brain; in addition to the mRNA, the antisense strand of zfh-5 is also expressed in the developing brain, in a manner complementary to the expression of zfh-5 mRNA. Although most neurons express zfh-5 mRNA soon after their final mitosis, several types of neurons, such as the pyramidal and granule cells in the hippocampus, express the zfh-5 antisense RNA prior to the mRNA expression. Using gene-targeting approach, we showed that this antisense RNA has a negative regulatory role on the expression of zfh-5 mRNA. These observations suggest that, in specific types of neurons, the expression of zfh-5 is additionally regulated by a mechanism depending on this antisense RNA.

The intracellular distribution of 10Sa RNA in Escherichia coli was investigated in cell extracts. Northern hybridization revealed that a large fraction of 10Sa RNA cosediments with 70S ribosomes. When 70S ribosomes were dissociated into 50S and 30S subunits in the presence of low levels of Mg2+ ions, almost all of the 10Sa RNA disappeared from both subunits. The extent of the association of the 10Sa RNA with ribosomes was much enhanced during the growth phase of the cells. These results suggest the possibility that 10Sa RNA might function on the ribosomes in E. coli cells.

Homeobox genes are important in the regulation of outgrowth and pattern formation during limb development. It is likely that homeobox genes play an equally important role during limb regeneration. We have isolated and identified 17 different homeobox-containing genes expressed by cells of regenerating axolotl limbs. Of these, nearly half of the clones represent genes belonging to the HoxA complex, which are thought to be involved in pattern formation along the proximal-distal limb axis. In this paper we report on the expression patterns of two 5′ members of this complex, HoxA13 and HoxA9. These genes are expressed in cells of developing limb buds and regenerating blastemas. The pattern of expression in developing axolotl limb buds is comparable to that in mouse and chick limb buds; the expression domain of HoxA13 is more distally restricted than that of HoxA9. As in developing mouse and chick limbs, HoxA13 likely functions in the specification of distal limb structures, and HoxA9 in the specification of more proximal structures. In contrast, during regeneration, HoxA13 and HoxA9 do not follow the rule of spatial colinearity observed in developing limbs. Instead, both genes are initially expressed in the same population of stump cells, giving them a distal Hox code regardless of the level of amputation. In addition, both are reexpressed within 24 hours after amputation, suggesting that reexpression may be synchronous rather than temporally colinear. Treatment with retinoic acid alters this Hox code to that of a more proximal region by the rapid and differential downregulation of HoxA13, at the same time that expression of HoxA9 is unaffected. HoxA reexpression occurs prior to blastema formation, 24–48 hours after amputation, and is an early molecular marker for dedifferentiation.

Zfhx2 (also known as zfh-5) encodes a transcription factor containing three homeobox domains and 18 Zn-finger motifs. We have reported that Zfhx2 mRNA is expressed mainly in differentiating neurons in the mouse brain and its expression level is negatively regulated by the antisense transcripts of Zfhx2. Although the expression profile of Zfhx2 suggests that ZFHX2 might have a role in a particular step of neuronal differentiation, the specific function of the gene has not been determined. We generated a Zfhx2-deficient mouse line and performed a comprehensive battery of behavioral tests to elucidate the function of ZFHX2. Homozygous Zfhx2-deficient mice showed several behavioral abnormalities, namely, hyperactivity, enhanced depression-like behaviors, and an aberrantly altered anxiety-like phenotype. These behavioral phenotypes suggest that ZFHX2 might play roles in controlling emotional aspects through the function of monoaminergic neurons where ZFHX2 is expressed. Moreover, considering their phenotypes, the Zfhx2-deficient mice may provide a novel model of human psychiatric disorders.

We have determined that 10Sa RNA (one of the small stable RNAs found in Escherichia coli) has an interesting structural feature: the 5' end and the 3' end of 10Sa RNA can be arranged in a structure that is equivalent to a half-molecule (acceptor stem and TFC stem-loop) of alanine tRNA of E. coli. Primer-extension analysis of 10Sa RNA extracted from a bacterial mutant with temperature-sensitive RNase P function revealed that the precursor to 10Sa RNA (pre-10Sa RNA) is folded into a pre-tRNA-like structure in vivo such that it can be cleaved by RNase P to generate the 5' end of the mature 10Sa RNA. The purified 10Sa RNA can be charged with alanine in vitro. Disruption of the gene encoding 10Sa RNA (ssrA) caused a reduction in the rate of cell growth, which was especially apparent at 45 degrees C, and a reduction in motility on semisolid agar. These phenotypic characteristics of the deletion strain (delta ssrA) allowed us to investigate the effects of some mutations in 10Sa RNA in vivo, although the exact function of 10Sa RNA still remains unclear. When the G.U pair (G3.U357) in 10Sa RNA, which may be equivalent to the determinant G.U pair of alanine tRNA, was changed to a G.A or G.C pair, the ability to complement the phenotypic mutations of the delta ssrA strain was lost. Furthermore, this inability to complement the mutant phenotypes that was caused by the substitution of the determinant bases by a G.A pair could be overcome by the introduction of a gene encoding alanyl-tRNA synthetase (alaS) on a multicopy plasmid. The evidence suggests that the proposed structural features of 10Sa RNA are indeed manifested in vivo.

It has been discovered that expression of promoter activity can be inhibited by visible light when specific fragments of E. coli DNA are inserted in a vector system designed to assay for promoter activity. These fragments have been located on regions of the E. coli chromosome to which no gene has been assigned to date. The effective wavelength of light that produces this phenomenon has been determined.

Protein D has previously been demonstrated to be associated with Escherichia coli ribosomes by the radical-free and highly reducing method of two-dimensional polyacrylamide gel electrophoresis. In this study, we show that protein D is exclusively present in the 30S ribosomal subunit and that its gene is located at 33.6 min on the E. coli genetic map, between ompC and sfcA . The gene consists of 45 codons, coding for a protein of 5,096 Da. The copy number of protein D per ribosomal particle varied during growth and increased from 0.1 in the exponential phase to 0.4 in the stationary phase. For these reasons, protein D was named SRA (stationary-phase-induced ribosome-associated) protein and its gene was named sra . The amount of SRA protein within the cell was found to be controlled mainly at the transcriptional level: its transcription increased rapidly upon entry into the stationary phase and was partly dependent on an alternative sigma factor (sigma S). In addition, global regulators, such as factor inversion stimulation (FIS), integration host factor (IHF), cyclic AMP, and ppGpp, were found to play a role either directly or indirectly in the transcription of sra in the stationary phase.

We have characterized a novel type of non-coding RNA which consists of tandem repeats of similar sequences, approximately 0.9 kb in size. This RNA, termed Bsr (brain specific repetitive) RNA, is encoded at a single locus (6 q31-->q32) in the rat genome, where 100 to 150 copies of the 0.9 kb sequences are repeated in tandem. Bsr RNA is preferentially expressed in the rat central nervous system (CNS), especially in phylogenetically old structures, such as the pareo- and archicortex, amygdala, thalamus and hypothalamus. In the developing brains, Bsr RNA is expressed in the subsets of differentiating cells but not in proliferating cells. Despite the finding that Bsr RNA appears to be conserved only among the Rattus species, the specific expression pattern of Bsr RNA suggests that it might have some role in the rat CNS.

The tRNAThr2 isoacceptor of E. coli has a G-A mismatch at positions 27–43. When the anticodon of this tRNA was converted to an amber anticodon (CUA), this tRNA showed suppressor activity in E. coli. Moreover, introduction of the base pair (G-C or U-A) at positions 27–43 of this suppressor tRNA reduced its suppressor activity. These results indicate that the G27-A43 mismatch is necessary for full function of tRNAThr2.

The chloroplast gene rps12 for ribosomal protein S12 in a liverwort, Marchantia polymorpha, is split into three exons by two introns, one of which (intron 1) is discontinuous. Exon 1 of rps12 for the N-terminal portion of the S12 protein is far from exons 2 and 3 for the C-terminal portion on the opposite DNA strand. S1-nuclease protection analysis and Northern hybridization with RNA isolated from the liverwort chloroplasts showed that: (i) the exons 1 and 2-3 of the rps12 gene with the neighboring genes were transcribed separately, (ii) the trans-splicing of intron 1 occurred after the processing of two primary transcripts to two pre-mRNAs, and (iii) there was no particular order for the splicing of intron 1 (trans) and intron 2 (cis) in the rps12 gene. We propose a bimolecular interaction model for trans-splicing by assuming that intermolecular base pairings between two pre-mRNAs result in the formation of the structure typical of group II introns except for disruption in the loop III region. This structure could be constructed in intron 1 of tobacco rps12 gene.