The priB gene of Klebsiella pneumoniae encodes a 104-amino acid protein that is similar in structure and function to Escherichia coli PriB

Primosome protein PriB is a single-stranded DNA-binding protein that serves as an accessory factor for PriA helicase-catalyzed origin-independent reinitiation of DNA replication in bacteria. A recent report describes the identification of a novel PriB protein in Klebsiella pneumoniae that is significantly shorter than most sequenced PriB homologs. The K. pneumoniae PriB protein is proposed to comprise 55 amino acid residues, in contrast to E. coli PriB which comprises 104 amino acid residues and has a length that is typical of most sequenced PriB homologs. Here, we report results of a sequence analysis that suggests that the priB gene of K. pneumoniae encodes a 104-amino acid PriB protein, akin to its E. coli counterpart. Furthermore, we have cloned the K. pneumoniae priB gene and purified the 104-amino acid K. pneumoniae PriB protein. Gel filtration experiments reveal that the K. pneumoniae PriB protein is a dimer, and equilibrium DNA binding experiments demonstrate that K. pneumoniae PriB's single-stranded DNA-binding activity is similar to that of E. coli PriB. These results indicate that the PriB homolog of K. pneumoniae is similar in structure and in function to that of E. coli.     

The amphotropic murine leukemia virus receptor gene encodes a 71-kilodalton protein that is induced by phosphate depletion.

The amphotropic murine leukemia virus (MuLV) can infect cells from a number of mammals, including humans, via its specific receptor. Basic knowledge of amphotropic MuLV receptor expression is likely to be useful in the development and improvement of gene therapy protocols based on amphotropic-pseudotyped vectors. To investigate the expression of the human receptor for the amphotropic MuLV (GLVR-2, newly termed Pit2), we determined its mRNA levels in several cell lines and found them to vary significantly. Induction of increased levels of mRNA after removal of phosphate from the media was observed in two osteosarcoma cell lines. The increase in GLVR-2 mRNA resulted in a concomitant rise in the levels of a 71-kDa protein specifically recognized by affinity-purified antibodies against GLVR-2. Using these antibodies, we were able to confirm the intracellular topology of the large hydrophilic domain between the proposed sixth and seventh transmembrane domains of the GLVR-2 protein. This assignment is in agreement with the fourth extracellular loop being outside the cell, consistent with the proposal that the fourth extracellular loop of GLVR-2 contains the envelope binding site.     

A gene located at 71F in Drosophila melanogaster encodes a novel zinc-finger protein that interacts with protein kinase CK2

Atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP) are two hormones produced and secreted by the heart to control blood pressure, body fluid homeostasis and electrolyte balance. Each peptide binds to a common family of 3 receptors (GC-A, GC-B and C-receptor) with varying degrees of affinity. The proANP gene disrupted mouse model provides an excellent opportunity to examine the regulation and expression of BNP in the absence of ANP. A new radioimmunoassay (RIA) was developed in order to measure mouse BNP peptide levels in the plasma, atrium and ventricle of the mouse. A detection limit of 3–6 pg/tube was achieved by this assay. Results show that plasma and ventricular level of BNP were unchanged among the three genotypes of mice. However, a significant decrease in the BNP level was noted in the atrium. The homozygous mutant (ANP–/–) had undetectable levels of BNP in the atrium, while the heterozygous (ANP+/–) and wild-type (ANP+/+) mice had 430 and 910 pg/mg in the atrium, respectively. Northern Blot analysis shows the ANP–/– mice has a 40% reduction of BNP mRNA level in the atrium and a 5-fold increase in the ventricle as compared with that of the ANP+/+ mouse. Our data suggest that there is a compensatory response of BNP expression to proANP gene disruption. Despite the changes in the atrial and ventricular tissue mRNA and peptide levels, the plasma BNP level remains unaltered in the ANP–/– mice. We conclude that the inability of BNP to completely compensate for the lack of ANP eventually leads to chronic hypertension in the proANP gene disrupted mice.     

The mouse Cebp gene encoding a DNA-binding protein is polymorphic and is located on chromosome 7

The mouse gene Cebp, encoding the DNA-binding protein C/EBP, has been localized to the proximal region of chromosome 7 by determining the strain distribution patterns of a restriction fragment length polymorphism among the BXD and AKXL recombinant inbred mouse lines.     

A retrotransposon-derived gene, PEG10, is a novel imprinted gene located on human chromosome 7q21

A novel paternally expressed imprinted gene, PEG10 (Paternally Expressed 10), was identified on human chromosome 7q21. PEG10 is located near the SGCE (Sarcoglycan epsilon) gene, whose mouse homologue was recently shown to be imprinted. Therefore, it is highly possible that a new imprinted gene cluster exists on human chromosome 7q21. Analysis of two predicted open reading frames (ORF1 and ORF2) revealed that ORF1 and ORF2 have homology to the gag and pol proteins of some vertebrate retrotransposons, respectively. These data suggest that PEG10 is derived from a retrotransposon that was previously integrated into the mammalian genome. PEG10 is likely to be essential for understanding how exogenous genes become imprinted.     

The segment polarity gene armadillo encodes a functionally modular protein that is the Drosophila homolog of human plakoglobin

The Drosophila segment polarity gene armadillo is required for pattern formation within embryonic segments and imaginal discs. We have found that armadillo is highly conserved during evolution; it is 63% identical to human plakoglobin, a protein found in adhesive junctions joining epithelial and other cells. We have examined arm protein localization in a number of larval tissues and found that arm protein accumulation within cells shares many features with the accumulation of plakoglobin. We have compared the phenotype and molecular lesions responsible for the different arm mutations. Surprisingly, severely truncated proteins retain some function; the degree of function is strictly correlated with the length of the truncated protein, suggesting that the internally repetitive arm protein is modular in function. We present a possible model for the cellular role of arm.     

The gene for histone RNA hairpin binding protein is located on human chromosome 4 and encodes a novel type of RNA binding protein.

The hairpin structure at the 3' end of animal histone mRNAs controls histone RNA 3' processing, nucleocytoplasmic transport, translation and stability of histone mRNA. Functionally overlapping, if not identical, proteins binding to the histone RNA hairpin have been identified in nuclear and polysomal extracts. Our own results indicated that these hairpin binding proteins (HBPs) bind their target RNA as monomers and that the resulting ribonucleoprotein complexes are extremely stable. These features prompted us to select for HBP-encoding human cDNAs by RNA-mediated three-hybrid selection in Saccharomyces cerevesiae. Whole cell extract from one selected clone contained a Gal4 fusion protein that interacted with histone hairpin RNA in a sequence- and structure-specific manner similar to a fraction enriched for bovine HBP, indicating that the cDNA encoded HBP. DNA sequence analysis revealed that the coding sequence did not contain any known RNA binding motifs. The HBP gene is composed of eight exons covering 19.5 kb on the short arm of chromosome 4. Translation of the HBP open reading frame in vitro produced a 43 kDa protein with RNA binding specificity identical to murine or bovine HBP. In addition, recombinant HBP expressed in S. cerevisiae was functional in histone pre-mRNA processing, confirming that we have indeed identified the human HBP gene.     

The murine endogenous retrovirus MIA14 encodes an active aspartic proteinase that is functionally similar to proteinases from D-type retroviruses

Murine intracisternal A-type particles (IAPs) are endogenous retroviruses showing sequence homologies to B/D- and avian C-type retroviruses and a gene expression strategy similar to that of D-type retroviruses. These viruses form immature particles in the endoplasmic reticulum and do not release extracellular virions, but are competent for retrotransposition within the virus-producing cell. It had been assumed that lack of polyprotein processing and maturation is due to a defect in the viral proteinase (PR), but recent experiments have shown that polyprotein processing occurs when assembly of the mouse IAP MIA14 is artificially directed to the plasma membrane. We have expressed and purified recombinant MIA14 PR and show that it undergoes N- and C-terminal autoprocessing at defined sites. Using peptide cleavage and inhibition assays and in vitro cleavage of recombinant HIV-1 and MIA14 Gag polyproteins, we show that MIA14 PR is a catalytically competent enzyme comparable in its efficiency to PRs from type D exogenous retroviruses. MIA14 PR is related to the PR of Mason-Pfizer monkey virus both functionally and with respect to its expression strategy, and is distinct from HIV-1 PR with respect to substrate specificity and catalytic efficiency. These findings reveal a functional and possibly evolutionary relationship between MIA14 and D-type retroviruses and imply that a functional PR may be relevant for intracellular retrotransposition even in the case of an endogenous retrovirus that does not produce extracellular virus.     

A prostate-derived cDNA that is mapped to human chromosome 19 encodes a novel protein

The epithelial cells of prostate gland secrete various secretory products that play an important role in the growth and differentiation of prostate gland. These secretory products have also been implicated in neuroendocrine differentiation of benign prostatic hyperplasia and prostate malignancy. We have cloned a prostate-derived cDNA encoding a novel protein with a predicted molecular weight of 78 kDa (P(78)), and precisely mapped the cDNA sequence to chromosome 19. The P(78) gene has a complex genomic structure with 18 exons and 17 introns. The P(78) contains two conserved structural domains with limited similarity to domain D of synapsin I. The P(78) mRNA was expressed in various human cell lines. Western blot analysis using antibody specific for the P(78) revealed the presence of the P(78) protein in the prostate cancer cell lines with much lower level in metastatic prostate cancer cell lines compared to that in a primary prostate cancer cell line.     

The yes-related cellular gene lyn encodes a possible tyrosine kinase similar to p56lck.

With v-yes DNA as the probe, a human cDNA library made from placental RNA was screened under relaxed conditions, and DNA clones derived from a novel genetic locus, termed lyn, were obtained. Nucleotide sequencing revealed that lyn could encode a novel tyrosine kinase that was very similar to mouse T-lymphocyte-specific tyrosine kinase p56lck and the v-yes protein as well as to the gene products of v-fgr and v-src. Northern hybridization analysis revealed that a 3.2-kilobase lyn mRNA was expressed in a variety of tissues of the human fetus. The pattern of lyn mRNA expression was different from those of related genes, such as yes and syn. Hybridization analysis of DNA from sorted chromosomes showed that the lyn gene is located on human chromosome 8 q13-qter.     

The pavA gene of Streptococcus pneumoniae encodes a fibronectin-binding protein that is essential for virulence

Streptococcus pneumoniae colonizes the nasopharynx in up to 40% of healthy subjects, and is a leading cause of middle ear infections (otitis media), meningitis and pneumonia. Pneumococci adhere to glycosidic receptors on epithelial cells and to immobilized fibronectin, but the bacterial adhesins mediating these reactions are largely uncharacterized. In this report we describe a novel pneumococcal protein PavA, which binds fibronectin and is associated with pneumococcal adhesion and virulence. The pavA gene, present in 64 independent isolates of S. pneumoniae tested, encodes a 551 amino acid residue polypeptide with 67% identical amino acid sequence to Fbp54 protein in Streptococcus pyogenes. PavA localized to the pneumococcal cell outer surface, as demonstrated by immunoelectron microscopy, despite lack of conventional secretory or cell-surface anchorage signals within the primary sequence. Full-length recombinant PavA polypeptide bound to immobilized human fibronectin in preference to fluid-phase fibronectin, in a heparin-sensitive interaction, and blocked binding of wild-type pneumococcal cells to fibronectin. However, a C-terminally truncated PavA' polypeptide (362 aa residues) failed to bind fibronectin or block pneumococcal cell adhesion. Expression of pavA in Enterococcus faecalis JH2-2 conferred > sixfold increased cell adhesion levels to fibronectin over control JH2-2 cells. Isogenic mutants of S. pneumoniae, either abrogated in PavA expression or producing a 42 kDa C-terminally truncated protein, showed up to 50% reduced binding to immobilized fibronectin. Inactivation of pavA had no effects on growth rate, cell morphology, cell-surface physico-chemical properties, production of pneumolysin, autolysin, or surface proteins PspA and PsaA. Isogenic pavA mutants of encapsulated S. pneumoniae D39 were approximately 104-fold attenuated in virulence in the mouse sepsis model. These results provide evidence that PavA fibronectin-binding protein plays a direct role in the pathogenesis of pneumococcal infections.     

The human gene AHNAK encodes a large phosphoprotein located primarily in the nucleus

AHNAK is a newly identified human gene notable for the exceptional size (c.a. 700 kD) and structure of its product, and for the repression of its expression in human neuroblastoma cells. Here we report the identification and partial characterization of the protein encoded by AHNAK. The protein is located principally (but not exclusively) in the nucleus and is phosphorylated on both serine and threonine. The abundance of the protein increases appreciably when cells withdraw from the division cycle, in response to either withdrawal of serum (fibroblasts) or differentiation (neuroblastoma cells). By contrast, the amount of phosphorylation appears to diminish in those settings. The considerable abundance and conjectured fibrous structure of AHNAK protein suggest a role in cytoarchitecture, but no function can yet be discerned.     

PHH1, a novel gene from Arabidopsis thaliana that encodes a protein similar to plant blue-light photoreceptors and microbial photolyases

A cDNA from Arabidopsis thaliana similar to microbial photolyase genes, and designated AT-PHH1, was isolated using a photolyase-like cDNA from Sinapsis alba (SA-PHR1) as a probe. Multiple isolations yielded only PHH1 cDNAs, and a few blue-light-receptor CRY1 (HY4) cDNAs (also similar to microbial photolyase genes), suggesting the absence of any other highly similar Arabidopsis genes. The AT-PHH1 and SA-PHR1 cDNA sequences predict 89% identity at the protein level, except for an AT-PHH1 C-terminal extension (111 amino acids), also not seen in microbial photolyases. AT-PHH1 and CRY1 show less similarity (54% protein identity), including respective C-terminal extensions that are themselves mostly dissimilar. Analysis of fifteen AT-PHH1 genomic isolates reveals a single gene, with three introns in the coding sequence and one in the 5′-untranslated leader. Full-length AT-PHH1, and both AT-PHH1 and AT-PHH1ΔC-513 (truncated to be approximately the size of microbial photolyase genes) cDNAs, were overexpressed, respectively, in yeast and Escherichia coli mutants hypersensitive to ultraviolet light. The absence of significant effects on resistance suggests either that any putative AT-PHH1 DNA repair activity requires cofactors/chromophores not present in yeast or E. coli, or that AT-PHH1 encodes a blue-light/ultraviolet-A receptor rather than a DNA repair protein.