Both P1 and P2 protamine genes are expressed in mouse, hamster, and rat

To date, in mammals except for the mouse and human, only one protamine variant has been isolated from sperm. These mammalian protamines share amino acid sequence homology with mouse protamine 1 (mP1), the tyrosine-containing variant. Southern blot analysis of restriction enzyme digests of hamster and rat liver DNA reveals the presence of sequences homologous to mP1, and also to mouse protamine 2 (mP2) cDNAs. Northern blots of hamster and rat total testis RNA probed with mP2 cDNA confirm that the protamine 2 gene in these species is transcribed into two size classes of mRNA of approximately 830 and 700 nucleotides. However, the relative abundance of the rat and hamster protamine 2 mRNAs (rP2 and hP2) in total testis is approximately 50-fold lower and 2- to 5-fold lower, respectively, than the mouse protamine 2 mRNA. Northern blot analysis of hamster and rat testis polysome gradients demonstrates that although the amount of rP2 mRNA and hP2 mRNA is reduced, both are present on polysomes. The decreased expression of rat and hamster protamine 2 mRNA relative to their protamine 1 counterparts contrasts protamine expression in the mouse testis, where approximately equal amounts of mP1 and mP2 protamine mRNAs are present. These results suggest differential expression of the P1 and P2 protamine genes in three closely related mammals.     

Expression and genomic imprinting of DCN, PON2 and PEG3 genes in porcine placenta

Imprinted genes play vital roles in the placental development and fetal growth in eutherian mammals. DCN (decorin), PON2 (paraoxonase 2) and PEG3 (paternally expressed 3) genes have been identified as imprinted genes in the mouse. Here, we detected the imprinting status of three genes in the porcine placenta on DG90 (day 90 of gestation) and the expression differences in Yorkshire and Meishan placenta on DG26, DG55 and DG90. The results indicated that the DCN and PON2 genes were not imprinted genes, while the PEG3 gene showed paternal monoallelic expression in porcine placenta. The expression of the DCN gene increased from DG26 to DG90 in both Yorkshire and Meishan pig placenta. However, this gene expression was greater in Yorkshire than Meishan pig on DG55. The expression of the PON2 gene was greater in Meishan pig than that in Yorkshire on DG26 and DG90. The PEG3 gene expression was not affected by day of pregnancy or breed. Data from the present study contribute to function genomic of porcine placental development.     

The chloroplast-derived trnW and trnM-e genes are not expressed in Arabidopsis mitochondria

Depending on their genetic origin, plant mitochondrial tRNAs are classified into three categories: the "native" and "chloroplast-like" mitochondrial-encoded tRNAs and the imported nuclear-encoded tRNAs. The number and identity of tRNAs in each category change from one plant specie to another. As some plant mitochondrial trn genes were found to be not expressed, and as all Arabidopsis thaliana mitochondrial trn genes are known, we systematically tested the expression of A. thaliana mitochondrial trn genes. Both the "chloroplast-like" trnW and trnM-e genes were found to be not expressed. These exceptions are remarkable since trnW and trnM-e are expressed in the mitochondria of other land plants. Whereas we could not conclude which tRNA(Met) compensates the lack of expression of trnM-e, we showed that the cytosolic tRNA(Trp) is present in A. thaliana mitochondria, thus compensating the absence of expression of the mitochondrial-encoded trnW.     

Divergent fructokinase genes are differentially expressed in tomato.

Two cDNA clones (Frk1 and Frk2) encoding fructokinase (EC 2.7.1.4) were isolated from tomato (Lycopersicon esculentum). The Frk2 cDNA encoded a deduced protein of 328 amino acids that was more than 90% identical with a previously characterized potato (Solanum tuberosum) fructokinase. In contrast, the Frk1 cDNA encoded a deduced protein of 347 amino acids that shared only 55% amino acid identity with Frk2. Both deduced proteins possessed and ATP-binding motif and putative substrate recognition site sequences identified in bacterial fructokinases. The Frk1 cDNA was expressed in a mutant yeast (Saccharomyces cerevisiae) line, which lacks the ability to phosphorylate glucose and fructose and is unable to grow on glucose or fructose. Mutant cells expressing Frk1 were complemented to grow on fructose but not glucose, indicating that Frk1 phosphorylates fructose but not glucose, and this activity was verified in extracts of transformed yeast. The mRNA corresponding to Frk2 accumulated to high levels in young, developing tomato fruit, whereas the Frk1 mRNA accumulated to higher levels late in fruit development. The results indicate that fructokinase in tomato is encoded by two divergent genes, which exhibit a differential pattern of expression during fruit development.     

Identification and physical mapping of genes expressed in the corpus luteum in cattle

A representational difference analysis was performed to identify genes expressed in the corpus luteum of cattle. The corpus luteum is an ovarian structure that is essential for the establishment and maintenance of pregnancy. Knowledge of gene expression and function of corpus luteum will be important to improve fertility in humans and domestic animals. Housekeeping genes were removed from the corpus luteum representation (tester) using skeletal muscle as the subtracting agent (driver). A total of 80 clones of the final subtraction product were analysed by sequencing and 11 new bovine gene sequences were identified (pBTCL1-11). The sequences were mapped to segments of 10 different chromosomes using a somatic cell hybrid panel and a radiation hybrid panel. With one exception the locations are in agreement with published comparative maps of cattle and man. Expression in corpus luteum was verified by RT-PCR for all the 11 clones.     

Dickkopf genes are co-ordinately expressed in mesodermal lineages.

Dickkopf-1 (dkk-1) is member of a novel family of secreted proteins and functions in head induction during Xenopus embryogenesis, acting as a potent inhibitor of Wnt signalling. Here we report: (1) the isolation of two additional murine members of the dkk family, dkk-2 and dkk-3; and (2) analysis of adult and embryonic gene expression of mouse dkk-1,-2, and -3, Xenopus dkk-1 as well as chicken dkk-3. Comparative developmental analyses of the dkk-1, dkk-2 and dkk-3 in mice indicate that these genes are both temporally and spatially regulated. They define overlapping deep domains in mesenchymal lineages suggesting a co-ordinated mode of action. All dkks show distinct and elevated expression patterns in tissues that mediate epithelial- mesenchyme transformations suggesting that they may participate in heart, tooth, hair and whisker follicle, limb and bone induction. In the limb buds expression of these genes are found in regions of programmed cell death. In a given organ, dkk-1 tends to be the earliest member expressed. Comparison with Xenopus dkk-1 and chicken dkk-3 shows evolutionarily conserved expression patterns. Our observations indicate that dkk genes constitute a new family of secreted proteins that may mediate inductive interactions between epithelial and mesenchymal cells.     

The fliU and fliV genes are expressed as a single ORF in Salmonella choleraesuis

A DNA fragment carrying flagellar genes was cloned from Salmonella choleraesuis. Compared to the corresponding DNA fragment of Salmonella muenchen, this fragment contained three ORFs instead of four shown in S. muenchen. The DNA sequence data showed that there was an insertion of nucleotide C in the ORF of the S. choleraesuis fliU gene, which resulted in the disappearance of a termination codon downstream. The recombinant plasmid pFU11 containing the coding region of the fliU gene made by PCR on S. choleraesuis genomic DNA was constructed and expressed in Escherichia coli in the presence of IPTG. As expected, a 45 kDa protein band was observed on a SDS-PAGE gel, in contrast to two with each having about a half of the molecular weight. These results demonstrated that the DNA sequence encoding one protein (FliU) in S. choleraesuis corresponded to the DNA sequence encoding two proteins (FliU and FliV) in S. muenchen. The protein encoded by this single ORF might carry out the functions of two separated proteins by folding in such a way that its conformation could function like two interdependent protein subunits.     

Wheat Ec metallothionein genes. Like mammalian Zn2+ metallothionein genes, wheat Zn2+ metallothionein genes are conspicuously expressed during embryogenesis.

A cDNA library was prepared from the bulk mRNA of mature wheat embryos and screened with mixed 32P-labeled oligonucleotide probes that encoded parts of the partial amino-acid sequence for the Zn-containing Ec protein. Each DNA insert in 11 positives from a screen of 10(5) plaques encoded a 5' untranslated and a 3' untranslated region, in addition to an open reading frame (of 81 amino acids) which, in every case, corresponded to at least 56 of the 59 amino acids in the partial polypeptide sequence previously determined for the Ec protein. The three different mRNA sequences encoded in the cDNA probably correspond to single-copy genes in the A, B and D genomes of hexaploid wheat. A wheat genomic library was screened with 32P-labeled cDNA and gave a single positive in a screen of 5 x 10(5) plaques. A 3.1-kb genomic fragment (gf-3.1) was sequenced and a cap site for the encoded mRNA was determined by primer extension. The gf-3.1 sequence encodes an intronless mRNA for the Ec protein and contains appreciable amounts of 5' and 3' flanking sequences. In addition to a putative TATA box, two inverted-repeat sequences and one direct-repeat sequence, the 5' flank in gf-3.1 contains a sequence similar to the abscisic-acid-responsive element in other higher-plant genes but does not contain sequences similar to the metal-responsive elements in animal metallothionein genes. Consistent with these findings, RNA blotting shows that accumulation of Ec mRNA is abundant in immature embryos, undetectable in germinated embryos and can be induced by adding abscisic acid, but not by adding Zn2+ to the medium in which mature wheat embryos are germinated. The findings suggest that the wheat Ec metallothionein genes, like mammalian liver metallothionein genes, are conspicuously expressed during embryogenesis.