The RNA interacting domain but not the protein interacting domain is highly conserved in ribosomal protein P0

Protein P0 interacts with proteins P1alpha, P1beta, P2alpha, and P2beta, and forms the Saccharomyces cerevisiae ribosomal stalk. The capacity of RPP0 genes from Aspergillus fumigatus, Dictyostelium discoideum, Rattus norvegicus, Homo sapiens, and Leishmania infantum to complement the absence of the homologous gene has been tested. In S. cerevisiae W303dGP0, a strain containing standard amounts of the four P1/P2 protein types, all heterologous genes were functional except the one from L. infantum, some of them inducing an osmosensitive phenotype at 37 degrees C. The polymerizing activity and the elongation factor-dependent functions but not the peptide bond formation capacity is affected in the heterologous P0 containing ribosomes. The heterologous P0 proteins bind to the yeast ribosomes but the composition of the ribosomal stalk is altered. Only proteins P1alpha and P2beta are found in ribosomes carrying the A. fumigatus, R. norvegicus, and H. sapiens proteins. When the heterologous genes are expressed in a conditional null-P0 mutant whose ribosomes are totally deprived of P1/P2 proteins, none of the heterologous P0 proteins complemented the conditional phenotype. In contrast, chimeric P0 proteins made of different amino-terminal fragments from mammalian origin and the complementary carboxyl-terminal fragments from yeast allow W303dGP0 and D67dGP0 growth at restrictive conditions. These results indicate that while the P0 protein RNA-binding domain is functionally conserved in eukaryotes, the regions involved in protein-protein interactions with either the other stalk proteins or the elongation factors have notably evolved.     

Annexin A9 is a periplakin interacting partner in membrane-targeted cytoskeletal linker protein complexes

Periplakin regulates keratin organisation and participates in the assembly of epidermal cornified envelopes. A proteomic approach identified annexin A9 as a novel interacting partner for periplakin N-terminus. The presence of annexin A9 in complexes with periplakin was confirmed by immunoblotting of proteins immunoprecipitated by anti-HA or anti-annexin A9 antibodies. Both endogenous and GFP-tagged annexin A9 co-localise with endogenous periplakin and transfected periplakin N-terminus at MCF-7 cell borders and aggregate after Okadaic acid treatment. Annexin A9 and periplakin co-localise in the epidermis and annexin A9 is up-regulated in differentiating keratinocytes, but the epidermal annexin A9 expression does not require periplakin.Structured summary of protein interactionsAnnexin-9 physically interacts with Periplakin by anti bait co-immunoprecipitation (View interaction) Periplakin physically interacts with Annexin-9 by anti tag co-immunoprecipitation (View Interaction: 1, 2) Periplakin and Annexin-9 colocalize by fluorescence microscopy (View Interaction: 1, 2, 3) Keratin-8 and Periplakin colocalize by fluorescence microscopy (View interaction)     

The insect immune protein hemolin is expressed during oogenesis and embryogenesis

Hemolin is the most abundant bacteria-induced proteins in Hyalophora cecropia hemolymph. Its structural features, both at the protein and gene level, ascribe this molecule to the immunoglobulin gene superfamily (IgSF) with particular homology to neural cell adhesion molecules. An increasing number of evidence suggest a role in immune recognition and in cell adhesion events. Hemolin is also developmentally regulated as suggested by changes in its concentration during larval and pupal ecdysis (Trenczek, T., 1998. Endogenous defense mechanisms of insects. Zoology 101, 298-315; Lanz-Mendoza, H., Faye, I., 1999. Physiological aspects of the immunoglobulin superfamily in invertebrates. Dev. Comp. Immunol. 23, 359-374). In the present study the expression of hemolin was investigated in oogenesis and in early embryogenesis. Our results reveal that hemolin is expressed in follicles and in epidermal and neural tissues of embryos.     

Expression of the highly conserved RNA binding protein KOC in embryogenesis.

The human KOC gene which is highly expressed in cancer shows typical structural features of an RNA binding protein. We analyzed the temporal and spatial expression pattern of KOC in mouse embryos at different gestational ages. The expression of KOC seems to be ubiquitous at early stages. During advanced gestation highest KOC expression occurs in the gut, pancreas, kidney, and in the developing brain. The expression pattern of KOC was compared to its Xenopus homologue Vg1-RBP during frog development. Similar expression was found in these organs suggesting an important functional role of the homologous proteins in embryonic development.     

Bambi is coexpressed with Bmp-4 during mouse embryogenesis

Signaling of TGF-β superfamily members is tightly controlled by an elaborate network of regulators (for recent review see Trends Genet. 15 (1999) 3; Genes Dev. 14 (2000) 627). Recently, the transmembrane protein BAMBI (BMP and activin membrane-bound inhibitor) has been shown to interfere with Bmp and activin-like signaling by inhibiting Tgf-β type I receptor activation (Nature 401 (1999) 480). In striking contrast to other Bmp antagonists like noggin (Cell 86 (1996) 599) or chordin (Cell 86 (1996) 589), BAMBI is strictly coexpressed with Bmp-4 during early Xenopus embryogenesis. The grouping of genes according to their shared complex spatial expression pattern and their involvement in the same biological signaling pathway has been referred to as synexpression group. This concept facilitates prognoses about the roles of a group member with unknown function. Apparently, only a minority of genes is organized in synexpression groups and up to now they have mainly been described in yeast and Xenopus (for review see Nature 402 (1999) 483). In the frog, BAMBI is a member of the Bmp-4 synexpression group (Nature 401 (1999) 480). We identified two murine homologues of BAMBI one of which, named Bambi-ψ, is a pseudogene. We show that the spatiotemporal expression pattern of Bambi closely matches that of Bmp-4 during mouse embryonic development. Moreover, we show that Bambi expression is induced in mouse embryonic fibroblasts by Bmp-4. Hence, we provide first evidence for the existence of an evolutionarily conserved Bmp-4 synexpression group in mammals.     

A novel putative transmembrane protein, IZP6, is expressed in neural cells during embryogenesis

Gene trapping in mouse embryonic stem cells is an efficient method for identifying new genes and examining their functions. This method has been used in an effort to identify some novel genes involved in mouse development. In the present paper, one such gene named IZP6 is reported. Expression of the IZP6 gene, as monitored by beta-galactosidase expression in heterozygous mice, was detected in a developmentally regulated fashion: the expression pattern has two phases during the embryogenesis. In the first phase, from embryonic day 11.5 (E11.5) until E14.5, the reporter gene is mainly expressed in the forebrain. In the second phase, from E15.5 until birth, expression in the forebrain becomes weaker but is still observed in the olfactory bulb and the skin around the eyes, nose, limbs and tail. Thus, IZP6 gene expression changes from the central nervous system (the first phase) to the peripheral tissues (the second phase) during development. The IZP6 gene encodes a protein of 228 amino acids. Analysis of the secondary structure of the IZP6 protein revealed four hydrophobic regions, indicating that the IZP6 protein is a four transmembrane region protein. These results suggest that IZP6 is a transmembrane protein related to neurogenesis in the mouse.     

The polarity protein Par6 is coupled to the microtubule network during molluscan early embryogenesis

Cell polarity, which directs the orientation of asymmetric cell division and segregation of fate determinants, is a fundamental feature of development and differentiation. Regulators of polarity have been extensively studied, and the critical importance of the Par (partitioning-defective) complex as the polarity machinery is now recognized in a wide range of eukaryotic systems. The Par polarity module is evolutionarily conserved, but its mechanism and cooperating factors vary among different systems. Here we describe the cloning and characterization of a pond snail Lymnaea stagnalis homologue of partitioning-defective 6 (Lspar6). The protein product LsPar6 shows high affinity for microtubules and localizes to the mitotic apparatus during embryonic cell division. In vitro assays revealed direct binding of LsPar6 to tubulin and microtubules, which is the first evidence of the direct interaction between the two proteins. The interaction is mediated by two distinct regions of LsPar6 both located in the N-terminal half. Atypical PKC, a functional partner of Par6, was also found to localize to the mitotic spindle. These results suggest that the L. stagnalis Par complex employs the microtubule network in cell polarity processes during the early embryogenesis. Identical sequence and localization of LsPar6 for the dextral and the sinistral snails exclude the possibility of the gene being the primary determinant of handedness.     

A role for CITED2, a CBP/p300 interacting protein, in colon cancer cell invasion

A thorough understanding of histone acetyltransferase CBP/p300-mediated regulation of gene expression and cell growth is essential to identify mechanisms relevant to the development of histone deacetylase (HDAC) inhibitor-based preventive and therapeutic strategies. We found that knockdown of CBP/p300 interacting coactivator with glutamic acid/aspartic acid-rich tail 2 (CITED2) increased colon cancer cell invasiveness in vitro. Gene expression profiling revealed that CITED2 knockdown induced matrix metalloproteinase-13 (MMP-13) gene expression in colon cancer cells. Butyrate, a naturally occurring HDAC inhibitor, induced CITED2 expression and downregulated MMP-13 expression in RKO cells. Additionally, ectopic expression of CITED2 arrested RKO cell growth. Thus, CITED2 regulates colon cancer invasion and might be a target for HDAC inhibitor-based intervention of colon cancer.     

Cdx protein interaction with Hoxa5 regulatory sequences contributes to Hoxa5 regional expression along the axial skeleton

Hox gene functions are intimately linked to correct developmental expression of the genes. The identification of cis-acting regulatory sequences and their associated trans-acting factors constitutes a key step in deciphering the mechanisms underlying the correct positioning of the functional domain of Hox genes along the anterior-posterior axis. We have identified DNA elements driving Hoxa5 regionalized expression in mice, using the 2.1-kb mesodermal enhancer (MES) localized in Hoxa5 3' flanking sequences as a starting point. The MES sequence comprises regulatory elements targeting Hoxa5 expression in the limbs, the urogenital and gastrointestinal tracts, and the cervical-upper thoracic region of the prevertebral column. A 164-bp DNA fragment within the MES caudally restricts Hoxa5 expression at the level of prevertebra 10, corresponding to the posterior limit of its functional domain. Cdx proteins directly bind to this element in vitro via two conserved sites. Preventing Cdx binding by mutating the sites causes caudal expansion of the transgene expression domain. Of all three murine Cdx proteins that bind this element in vitro, Cdx4 has emerged as a potential regional posterior repressor of Hoxa5 expression. The restrictive control provided by Cdx interactions with Hoxa5 regulatory sequences may be one of the critical events in cervicothoracic axial specification.     

Localization of the fushi tarazu protein during Drosophila embryogenesis

The fushi tarazu (ftz) gene of Drosophila acts early in embryogenesis to regulate body segmentation. The localization of the ftz protein product in embryos was examined using indirect immunofluorescence microscopy. Antibodies were prepared against a β-galactosidase-ftz hybrid protein made in E. coli. The ftz protein was first detectable in blastoderm-stage embryos as seven stripes of nuclei encircling the embryos transversely. The stripes persist through the early events of gastrulation, but disappear before overt segmentation is visible. The ftz protein is expressed a second time in some nuclei of the developing nervous system. In contrast to the early pattern, at the later stage, ftz is expressed in each of fifteen metameric subunits of the embryo.     

A deubiquitinating activity is conserved in the large tegument protein of the herpesviridae

The largest tegument protein of herpes simplex virus 1 (HSV-1), UL36, contains a novel deubiquitinating activity embedded in it. All members of the Herpesviridae contain a homologue of HSV-1 UL36, the N-terminal segments of which show perfect conservation of those residues implicated in catalysis. For murine cytomegalovirus and Epstein-Barr virus, chosen as representatives of the beta- and gammaherpesvirus subfamilies, respectively, we here show that the homologous modules indeed display deubiquitinating activity in vitro. The conservation of this activity throughout all subfamilies is indicative of an important, if not essential, function.     

TUBA8: A new tissue-specific isoform of alpha-tubulin that is highly conserved in human and mouse

We have cloned and sequenced a cDNA from a human adult skeletal muscle cDNA library, encoding for a novel isoform of alpha-tubulin (tuba8) that is preferentially expressed in heart, skeletal muscle, and testis. A genomic DNA sequence from the chromosomal region 22q11 allowed us to determine the complete structure of the TUBA8 gene that mirrors the canonical exon/intron organization of the vertebrate alpha-tubulin genes. We also cloned and sequenced the cDNA of its murine homologue (MMU-TUBA8). The latter encodes for a protein that differs from its human counterpart in only three amino acids, revealing an extreme rate of conservation that is even extended to both the 3' and 5' UTRs of the mRNAs. Sequence comparison of these novel isoforms with other known alpha tubulins shows that tuba8 is the most divergent member of the mammalian alpha-tubulin family. The sequence peculiarity of the human and murine tuba8 strongly suggests that they might have functional significance and, according to the multi-tubulin hypothesis, that they might play specific functional roles in the cell cytoskeleton.