Unusual abundance of vertebrate 3-phosphate dehydrogenase pseudogenes
ABSTRACT Only one gene coding for glyceraldehyde 3-phosphate dehydrogenase (GAPDH, EC 184.108.40.206), a key enzyme in the control of glycolysis, is known to be functional in man, mouse, rat and chicken. The gene has been localized to chromosome 12 in human and chromosome 6 in mouse. Only a single mRNA species has been found in chicken and rat. However, analysis of genomic DNA blots of various species with a cloned GAPDH cDNA probe has revealed large differences in the level of reiteration, ranging from one to over 200 copies. On this basis, we have grouped these organisms into three classes according to the number of GAPDH-related sequences they contain; one class with a unique representation (chicken), another class of relatively low reiteration (10-30 copies in man, hare, guinea-pig and hamster) and a third class of high reiteration (greater than 200 copies in mouse and rat). The third class represents the first reported occurrence of such an extreme number of pseudogenes related to an enzyme-coding gene and suggests that a dramatic amplification event took place between 15 and 25 million years ago.
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- "Comparative genomics also demonstrates that the Smcp and Prm3 genes have not created retroposons in 16 and 12 species of mammals, respectively ( , Bagarova and Kleene, unpublished ) and Southern blots indicate that a variety of genes that are expressed specifically in meiotic and haploid spermatogenic cells are single copy . The observation that the somatic glyceraldehyde-3′- phosphate dehydrogenase (GAPDH) mRNA has generated ~200 processed pseudogenes, while the spermatid-specific GAPDH mRNA has generated none   , provides a striking example of the deficit in creation of retroposons from mRNAs that are expressed in haploid spermatogenic cells. Knockouts of the Mili2 and Dnmt3L genes reveal mechanisms that repress expression of LINE-1 retroposons in meiotic spermatogenic cells  , which probably suppress the creation of retroposons from cellular mRNAs . "
ABSTRACT: The comparative genomics of the Odf1, Prm1, Prm2, Tnp1, and Tnp2 genes in 13-21 diverse mammalian species reveals striking similarities and differences in the sequences that probably function in the transcriptional and translational regulation of gene expression in haploid spermatogenic cells, spermatids. The 5' flanking regions contain putative TATA boxes and cAMP-response elements (CREs), but the TATA boxes and CREs exhibit gene-specific sequences, and an overwhelming majority of CREs differ from the consensus sequence. The 5' and 3' UTRs contain highly conserved gene-specific sequences including canonical and noncanonical poly(A) signals and a suboptimal context for the Tnp2 translation initiation codon. The conservation of the 5' UTR is unexpected because mRNA translation in spermatids is thought to be regulated primarily by the 3' UTR. Finally, all of the genes contain a single intron, implying that retroposons are rarely created from mRNAs that are expressed in spermatids.Genomics 09/2008; 92(2):101-6. DOI:10.1016/j.ygeno.2008.05.001 · 2.79 Impact Factor
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- "A particularly illustrative example is glyceraldehyde-3-phosphate dehydrogenase (GAPDH), which seems to have evolved into a multifunctional protein through incremental changes in amino acid sequence that were neutral with respect to its housekeeping function but that endowed the altered versions with new, unrelated functions. In this regard, the NDK family may have provided an even richer source of novel functions because although GAPDH is encoded by a single gene (Piechaczyk et al., 1984), the NDK family consists of eight distinct genes (Nm23H1–H8), each differing in tissue distribution and subcellular localization. Nearly all NDK isoforms contain the set of highly conserved residues critical for nucleotide binding and hydrolysis, but these common elements are interspersed with variable regions in the different isoforms, leading to the speculation that the variable regions may be responsible for functions unrelated to nucleotide synthesis (Lacombe et al., 2000). "
ABSTRACT: The cytosolic coat protein complex II (COPII) mediates vesicle formation from the endoplasmic reticulum (ER) and is essential for ER-to-Golgi trafficking. The minimal machinery for COPII assembly is well established. However, additional factors may regulate the process in mammalian cells. Here, a morphological COPII assembly assay using purified COPII proteins and digitonin-permeabilized cells has been applied to demonstrate a role for a novel component of the COPII assembly pathway. The factor was purified and identified by mass spectrometry as Nm23H2, one of eight isoforms of nucleoside diphosphate kinase in mammalian cells. Importantly, recombinant Nm23H2, as well as a catalytically inactive version, promoted COPII assembly in vitro, suggesting a noncatalytic role for Nm23H2. Consistent with a function for Nm23H2 in ER export, Nm23H2 localized to a reticular network that also stained for the ER marker calnexin. Finally, an in vivo role for Nm23H2 in COPII assembly was confirmed by isoform-specific knockdown of Nm23H2 by using short interfering RNA. Knockdown of Nm23H2, but not its most closely related isoform Nm23H1, resulted in diminished COPII assembly at steady state and reduced kinetics of ER export. These results strongly suggest a previously unappreciated role for Nm23H2 in mammalian ER export.Molecular Biology of the Cell 03/2005; 16(2):835-48. DOI:10.1091/mbc.E04-09-0785 · 4.55 Impact Factor
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- "The purified extended products were analyzed on 7% polyacrylamide denaturing gels and exposed to Dupont reflection film. genase gene (GAPDH) (Karin et al., 1984; Piechaczyk et al., 1984; Koropatnick and Duerksen, 1987). In contrast with what is observed with the MMTV promoter in these cells, the MT mRNA levels were elevated at 24 and 48 h of hormone treatment (Figure 6A, compare lanes 1–4). "
ABSTRACT: Glucocorticoids rapidly induce transcription from the mouse mammary tumour virus (MMTV) promoter via a glucocorticoid receptor (GR)-mediated chromatin disruption event. This remodelling of chromatin is transient such that upon prolonged exposure to hormone the promoter becomes refractory to glucocorticoids. We demonstrate that this refractory state requires the continual presence of hormone and can be reversed by its removal. Our experiments show that the promoter is inactivated via a mechanism whereby histone H1 is dephosphorylated in response to glucocorticoids. Removal of glucocorticoids results in the rephosphorylation of histone H1 and the reacquisition of transcriptional competence by the promoter. This response is specific for the MMTV promoter assembled as chromatin and is not observed for another inducible gene or transiently transfected MMTV DNA. Finally, we demonstrate that H1 on the MMTV promoter is dephosphorylated when the promoter is unresponsive to glucocorticoids. These studies indicate that phosphorylated H1 is intimately linked with the GR-mediated disruption of MMTV chromatin in vivo.The EMBO Journal 04/1998; 17(5):1454-66. DOI:10.1093/emboj/17.5.1454 · 10.75 Impact Factor