RESUMO
Inbred mice are used to investigate many aspects of human physiology, including susceptibility to disease and response to therapies. Despite increasing evidence that the composition and function of the murine intestinal microbiota can substantially influence a broad range of experimental outcomes, relatively little is known about microbiome dynamics within experimental mouse populations. We investigated changes in the intestinal microbiome between C57BL/6J mice spanning six generations (assessed at generations 1, 2, 3, and 6), following their introduction to a stringently controlled facility. Fecal microbiota composition and function were assessed by 16S rRNA gene amplicon sequencing and liquid chromatography mass spectrometry, respectively. Significant divergence of the intestinal microbiota between founder and second generation mice, as well as continuing inter-generational variance, was observed. Bacterial taxa whose relative abundance changed significantly through time included Akkermansia, Turicibacter, and Bifidobacterium (p < 0.05), all of which are recognized as having the potential to substantially influence host physiology. Shifts in microbiota composition were mirrored by corresponding differences in the fecal metabolome (r = 0.57, p = 0.0001), with notable differences in levels of tryptophan pathway metabolites and amino acids, including glutamine, glutamate and aspartate. We related the magnitude of changes in the intestinal microbiota and metabolome characteristics during acclimation to those observed between populations housed in separate facilities, which differed in regards to husbandry, barrier conditions and dietary intake. The microbiome variance reported here has implications for experimental reproducibility, and as a consequence, experimental design and the interpretation of research outcomes across wide range of contexts.
RESUMO
Autism spectrum disorder (ASD) is a common neurodevelopmental condition characterized by marked genetic heterogeneity. Recent studies of rare structural and sequence variants have identified hundreds of loci involved in ASD, but our knowledge of the overall genetic architecture and the underlying pathophysiological mechanisms remains incomplete. Glycine receptors (GlyRs) are ligand-gated chloride channels that mediate inhibitory neurotransmission in the adult nervous system but exert an excitatory action in immature neurons. GlyRs containing the α2 subunit are highly expressed in the embryonic brain, where they promote cortical interneuron migration and the generation of excitatory projection neurons. We previously identified a rare microdeletion of the X-linked gene GLRA2, encoding the GlyR α2 subunit, in a boy with autism. The microdeletion removes the terminal exons of the gene (GLRA2(Δex8-9)). Here, we sequenced 400 males with ASD and identified one de novo missense mutation, p.R153Q, absent from controls. In vitro functional analysis demonstrated that the GLRA2(Δex8)(-)(9) protein failed to localize to the cell membrane, while the R153Q mutation impaired surface expression and markedly reduced sensitivity to glycine. Very recently, an additional de novo missense mutation (p.N136S) was reported in a boy with ASD, and we show that this mutation also reduced cell-surface expression and glycine sensitivity. Targeted glra2 knockdown in zebrafish induced severe axon-branching defects, rescued by injection of wild type but not GLRA2(Δex8-9) or R153Q transcripts, providing further evidence for their loss-of-function effect. Glra2 knockout mice exhibited deficits in object recognition memory and impaired long-term potentiation in the prefrontal cortex. Taken together, these results implicate GLRA2 in non-syndromic ASD, unveil a novel role for GLRA2 in synaptic plasticity and learning and memory, and link altered glycinergic signaling to social and cognitive impairments.
Assuntos
Glicina/metabolismo , Receptores de Glicina/genética , Receptores de Glicina/metabolismo , Adolescente , Adulto , Animais , Transtorno do Espectro Autista/metabolismo , Transtorno Autístico/metabolismo , Criança , Pré-Escolar , Glicina/genética , Humanos , Potenciação de Longa Duração/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurogênese/efeitos dos fármacos , Plasticidade Neuronal/efeitos dos fármacos , Neurônios/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transmissão Sináptica/fisiologia , Peixe-ZebraRESUMO
Calpains are a family of related proteins, some of which have been shown to function as calcium-dependent cysteine proteases. CAPN1 and CAPN2, the most well characterized calpains, consist of a large (80 kDa) and a small (30 kDa) subunit. In mammals, 11 different paralogous genes encoding calpain large subunits have been identified. We report the identification of two further genes, CAPN13 and CAPN14, potentially encoding calpain large subunits. Radiation hybrid mapping localized both genes within a region mapped to 2p21-2p22. The CAPN13 mRNA exhibits a restricted tissue distribution with low levels of expression detected only in human testis and lung while CAPN14 mRNA could not be detected in any of the 76 tissues examined. Examination of the human genome sequence in the public and private consortia databases did not detect any further members of this gene family. Thus, there would seem to be 13 large subunit calpain genes in the human genome. Phylogenetic analysis reveals that the putative calpain large subunit proteins can be divided into three major groups. The 13 human large subunit genes and the single small subunit gene are located in eight syntenic groups on chromosomes 1, 2, 3, 6, 11, 15, 19 and X.
Assuntos
Calpaína/genética , Família Multigênica/genética , Sequência de Aminoácidos , Northern Blotting , Cromossomos Humanos Par 2/genética , Expressão Gênica , Ordem dos Genes , Humanos , Masculino , Dados de Sequência Molecular , Filogenia , Subunidades Proteicas , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Mapeamento de Híbridos Radioativos , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Distribuição TecidualRESUMO
The genetic steps governing development of the spleen are largely unknown. Absence of Hox11 in mice results in asplenia, but it is unclear how Hox11 exerts its effect on spleen development. To more precisely define Hox11's role in spleen morphogenesis, we have examined the fate of the developing spleen in Hox11(-/-) mice. Perturbation of spleen development begins between dE13 and dE13.5. Cells of the spleen anlage persist past this developmental stage as an unorganized rudiment between the stomach and the pancreas. They fail to proliferate, and haematopoietic cells do not colonize the rudiment. At later stages of embryonic development, the cells can be observed in the mesenchyme of the pancreas, also an expression site of Hox11. In Hox11-/-<-->+/+ chimaeras, spleens were devoid of Hox11(-/-) cells, indicating that the genetic defect is cell autonomous and not due to failure of the organ anlage to attract and retain haematopoietic cells. In -/-<-->+/+ chimaeric embryos, Hox11(-/-) cells were initially present in the spleen anlage. However, at dE13, a reorganization of the spleen occurred in the chimaeras and Hox11(-/-) cells were subsequently excluded from the spleen, suggesting that a change in the affinity for one of the spleen cells had occurred. These observations demonstrate that spleen development consists of genetically separable steps and that absence of Hox11 arrests spleen development at an early stage. The formation of the spleen primordium before the entry of haematopoietic cells does not require the activity of Hox11. However, subsequent differentiation of spleen precursor cells is dependent on the Hox11 gene.
Assuntos
Proteínas de Homeodomínio/metabolismo , Mesoderma/citologia , Proteínas Oncogênicas/metabolismo , Baço/embriologia , Animais , Agregação Celular , Diferenciação Celular , Quimera , Proteínas de Homeodomínio/genética , Camundongos , Camundongos Mutantes , Mórula , Proteínas Oncogênicas/genética , Células-TroncoAssuntos
Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Dedos de Zinco/genética , Sequência de Aminoácidos , Animais , Mapeamento Cromossômico , Cromossomos Humanos Par 19 , Clonagem Molecular , Etiquetas de Sequências Expressas , Regulação da Expressão Gênica no Desenvolvimento , Glicosilação , Humanos , Masculino , Camundongos , Dados de Sequência Molecular , Fosforilação , Homologia de Sequência de Aminoácidos , Testículo/fisiologiaRESUMO
We report the identification of mouse Capn12, a new member of the calpain large subunit gene family. It possesses potential protease and calcium-binding domains, features typical of the classical calpains. In situ hybridization and Northern blot analysis demonstrate that during the anagen phase of the hair cycle the cortex of the hair follicle is the major expression site of Capn12. The gene was sequenced in its entirety and consists of 21 exons spanning 13 kb with an exon-intron structure typical of the calpain gene family. The last exon of the mouse Actn4 gene overlaps the 3' end of Capn12 but in the opposite orientation. This overlap between the two genes is conserved in the human genome. Three versions of the Capn12 mRNA transcript were identified. They occur as a result of alternative splicing, and two of these encode a protein lacking the C-terminal calmodulin-like domain. Radiation hybrid mapping localized Capn12 to mouse chromosome 7, closely linked to a marker positioned at 10.4 cM. Refined mapping of Capn5, also previously localized to chromosome 7, indicated that it was not closely linked to Capn12, mapping tightly linked to a marker positioned at 48.5 cM.
Assuntos
Calpaína/genética , Genes/genética , Proteínas dos Microfilamentos , Actinina/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Northern Blotting , Mapeamento Cromossômico , DNA/química , DNA/genética , Éxons , Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Homologia de Genes , Células Híbridas , Hibridização In Situ , Íntrons , Camundongos , Camundongos Endogâmicos , Dados de Sequência Molecular , Filogenia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Pele/crescimento & desenvolvimento , Pele/metabolismoRESUMO
Mice deficient in Hox11 are asplenic. As Hoxll can function as a transcription factor, we examined the spatial and temporal mRNA expression patterns of Hox11 and a candidate target gene, the Wilm's tumor gene Wt1, in the developing spleen. Hox11 mRNA first appears at approximately dE10.5 in the dorsal mesogastrium while Wt1 mRNA is expressed from dE11.5, approximately 24 hours after Hox11 mRNA first appears. Wt1 mRNA was significantly reduced in the spleen anlage of Hox11-null mice suggesting that Wt1 acts downstream of Hox11 in a transcriptional cascade. Additionally, Hox11 protein is able to transactivate the WT1 promoter in a Hox11-null fibroblast cell line. As Wt1-null embryos have recently been reported to be asplenic, these findings suggest that Wt1 and Hox11 may be components common to a genetic hierarchy that is required for spleen development.
Assuntos
Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas de Homeodomínio/genética , Proteínas Oncogênicas/genética , Baço/anormalidades , Baço/embriologia , Fatores de Transcrição/genética , Animais , Linhagem Celular , Feminino , Genes Reporter , Hibridização In Situ , Luciferases/genética , Masculino , Camundongos , Camundongos Mutantes , Gravidez , Regiões Promotoras Genéticas/fisiologia , RNA Mensageiro/análise , Transcrição Gênica/fisiologia , Proteínas WT1RESUMO
Calpains are a family of related proteins, originally classified on the basis of their calcium dependence and protease activity. Here we report the mRNA expression patterns during mouse development of the recently identified Capn5, Capn6 and Capn11 genes. The major expression sites of Capn5 during embryogenesis are the developing thymus, sympathetic and dorsal root ganglia. Capn6 mRNA is exclusively expressed during embryogenesis predominantly in developing skeletal and heart muscle overlapping closely with Capn3 expression domains. Expression was also observed in specific cells of the lung, kidney and placenta and in various epithelial cell types where the Capn6 mRNA appeared to be localized within the cell to the basal and apical ends. Capn11 mRNA is restricted exclusively to spermatocytes and only during the later stages of meiosis.
Assuntos
Calpaína/genética , Regulação da Expressão Gênica no Desenvolvimento , Testículo/embriologia , Animais , Animais Recém-Nascidos , Calpaína/metabolismo , Masculino , Camundongos , Desenvolvimento Muscular , Músculo Esquelético/embriologia , Músculo Esquelético/crescimento & desenvolvimento , Especificidade de Órgãos , RNA MensageiroRESUMO
The molecular basis of the characteristic hair growth disorder in nude mice that carry a defective Whn transcription factor gene is unknown. A comparison of mRNA populations from wild-type and nude mice back skin by representational difference analysis revealed the absence of acidic hair keratin gene 3 (mHa3) mRNA in mutant mice. Whn and acidic hair keratin genes are co-expressed in hair follicles, nail forming regions and filiform papillae of the tongue: expression of the mHa3 gene is generally detectable about 1 day after Whn mRNA and rapidly ceases in its absence. Whn is strongly expressed during the anagen (growth) phase of the hair cycle in matrix, cortex and outer root sheath; its expression rapidly declines during catagen and is undetectable in telogen phases. In nude mice, low levels of mHa3 expression are maintained in nails and whisker follicles, whereas expression is completely absent in pelage hair follicles and filiform papillae. Thus, the nude phenotype represents the first example of an inherited skin disorder that is associated with the loss of expression rather than structural mutation of keratin genes. The distinct molecular difference between pelage and whisker follicles correlates with the improved mechanical stability of vibrissae in nude mice, implicating mHa3 as an important structural component of the hair shaft.
Assuntos
Diferenciação Celular/genética , Proteínas de Ligação a DNA/genética , Folículo Piloso/fisiologia , Queratinas/genética , Fatores de Transcrição/genética , Animais , Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição Forkhead , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Queratinas/metabolismo , Queratinas Específicas do Cabelo , Queratinas Tipo I , Camundongos , Camundongos Endogâmicos , Camundongos Mutantes , Camundongos Nus , Timo/embriologia , Fatores de Transcrição/metabolismoRESUMO
Calpains are a superfamily of related proteins, some of which have been shown to function as calcium-dependent cysteine proteases. In mammals, eight different calpains have been identified. We report the identification of a new mammalian calpain gene, CAPN11. The predicted protein possesses the features typical of calpains including potential protease and calcium-binding domains. The CAPN11 mRNA exhibits a highly restricted tissue distribution with highest levels present in testis. Radiation hybrid mapping localized the gene to human chromosome 6, within a region mapped to p12. Phylogenetic analysis suggests that, in mammals, the predicted CAPN11 protein is most closely related to CAPN1 and CAPN2. However, of the calpain sequences available, the predicted CAPN11 sequence exhibits greatest homology to the chicken micro/m calpain. Thus CAPN11 may be the human orthologue of micro/m calpain. The discovery of this new calpain emphasizes the complexity of the calpain family, with members being distinguished on the basis of protease activity, calcium dependence, and tissue expression.
Assuntos
Calpaína/genética , Sequência de Aminoácidos , Mapeamento Cromossômico , Cromossomos Humanos Par 6/genética , DNA Complementar/química , DNA Complementar/genética , Evolução Molecular , Feminino , Expressão Gênica , Humanos , Masculino , Dados de Sequência Molecular , Filogenia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Testículo/metabolismo , Distribuição TecidualRESUMO
We have studied changes in gene expression between normal and nude mouse skin. Using the method of representational difference analysis, a cDNA encoding a novel serine protease designated BSSP was cloned. Our results show that in skin this gene is predominantly expressed in the sebaceous gland of the hair follicle and in the distal part of the outer root sheath. Thus, BSSP is the first serine protease known to be expressed in the sebaceous gland. In nude mouse skin, this gene is overexpressed.
Assuntos
Cabelo/enzimologia , Calicreínas , Serina Endopeptidases/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA Complementar , Expressão Gênica , Camundongos , Camundongos Nus , Dados de Sequência Molecular , Serina Endopeptidases/metabolismo , Pele/metabolismoAssuntos
Calpaína/genética , Vertebrados/genética , Sequência de Aminoácidos , Animais , Sítios de Ligação , Northern Blotting , Cálcio/metabolismo , Calpaína/química , Calpaína/metabolismo , Camundongos , Dados de Sequência Molecular , Nematoides/genética , Filogenia , Homologia de Sequência de AminoácidosRESUMO
Helix-loop-helix proteins constitute a family of transcription factors with the potential to form homo- and hetero-dimers mediated by the helix-loop-helix domain. Oncogenic mutations in such genes can disrupt the equilibrium of protein-protein interactions in the affected cell. In order to assess the biological consequences of such mutations, the full complement of interacting proteins must be known. To identify proteins interacting with the basic-helix-loop-helix domain of the ubiquitously expressed E47 protein, a 'sandwich'-screening procedure was developed which distinguishes between homo- and hetero-oligomers, and specifically excludes the detection of complexes which cannot bind DNA. Nine distinct cDNAs were identified which encode proteins with apparent basic-helix-loop-helix domains, including a novel clone termed eip1 which is distantly related in the basic-helix-loop-helix domain to the Drosophila enhancer-of-split m7 protein. Using epitope-tagging, interaction of E47 basic-helix-loop-helix protein with the eip1 protein encoded by this novel cDNA was confirmed by immunoprecipitation experiments in COS7 cells. Interaction was also observed in the yeast two-hybrid system. Three cDNAs encoding proteins without basic-helix-loop-helix domains were also found to interact in the sandwich-expression screen. Interactions with human PARP and mouse replication factor 1a were confirmed using glutathione transferase-tagged cDNAs. A cDNA encoding part of the nucleolin protein sequence interacted with the E47 basic-helix-loop-helix only when fused to a beta-galactosidase tag.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Sequências Hélice-Alça-Hélice , Proteínas de Homeodomínio , Fatores de Transcrição/metabolismo , Sequência de Aminoácidos , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Clonagem Molecular/métodos , DNA Complementar/genética , Humanos , Substâncias Macromoleculares , Camundongos , Dados de Sequência Molecular , Testes de Precipitina , Ligação Proteica , Proteínas Recombinantes/metabolismo , Fatores de Transcrição TCF , Proteína 1 Semelhante ao Fator 7 de Transcrição , Fatores Estimuladores UpstreamRESUMO
The differentiation of primitive epithelial precursor cells in the thymic primordium into subcapsular, cortical, and medullary epithelial cells of the mature thymus requires the activity of the nude gene product Whn. Whn is also required for proper keratinization of the hair shaft. We determined the nucleotide sequence of a 58 kilobase region on mouse chromosome 11 that encompasses the mouse nude gene and part of the two neighboring genes, encoding a sodium/dicarboxylate co-transporter and the retinal protein 4. Using cross-hybridization, the human orthologue of the mouse nude gene was isolated. The human WHN protein also consists of 648 amino acids, 85% of which are identical to the mouse protein. Like the mouse gene, the human gene consists of eight coding exons and utilizes two alternative first exons in a tissue-specific fashion. Sequences upstream of the two alternative first exons display promoter activity in heterologous reporter assays. Whereas both promoters appear to be active in skin (albeit at different levels), only the most upstream element is active in the thymus, indicating that transcriptional activity of the whn gene is subject to complex regulation. Nucleotide sequence database comparisons reveal that among other winged-helix genes, the HTLF and HTLFL1 genes are most closely related to whn, although the exon/intron structure of the human HTLF gene in the DNA binding domain differs from that of whn.
Assuntos
Proteínas de Ligação a DNA/genética , Fatores de Transcrição/genética , Processamento Alternativo , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA Complementar , Proteínas de Ligação a DNA/química , Éxons , Fatores de Transcrição Forkhead , Genoma , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Homologia de Sequência de Aminoácidos , Pele/metabolismo , Timo/metabolismo , Timo/patologia , Fatores de Transcrição/químicaRESUMO
The HOX11 homeobox gene was identified via the translocation t(10;14) in T cell leukaemia. To determine the function of this gene in mice, null mutations were made using homologous recombination in ES cells to incorporate lacZ into the hox11 transcription unit. Production of beta-galactosidase from the recombinant hox11 allele in +/- mutants allowed identification of sites of hox11 expression which included the developing spleen. Newborn hox11 -/- mice exhibit asplenia. Spleen formation commences normally at E11.5 in hox11 -/- mutant embryos but the spleen anlage undergoes rapid and complete resorption between E12.5 and E13.5. Dying spleen cells exhibit molecular features of apoptosis, suggesting that programmed cell death is initiated at this stage of organ development in the absence of hox11 protein. Thus hox11 is not required to initiate spleen development but is essential for the survival of splenic precursors during organogenesis. This function for hox11 suggests that enhanced cell survival may result from the t(10;14) which activates HOX11 in T cell leukaemias, further strengthening the association between oncogene-induced cell survival and tumorigenesis.
Assuntos
Genes Homeobox , Proteínas de Homeodomínio/genética , Proteínas Oncogênicas/genética , Baço/embriologia , Animais , Apoptose/genética , Sequência de Bases , Expressão Gênica , Hibridização In Situ , Leucemia de Células T/genética , Camundongos , Camundongos Mutantes , Dados de Sequência Molecular , Morfogênese/genética , Mutagênese Sítio-Dirigida , Sondas de Oligonucleotídeos/genética , Baço/anormalidades , Baço/citologia , Translocação GenéticaRESUMO
The human homeobox gene HOX11 has been identified at the site of a chromosomal translocation in a subset of T-cell acute leukaemias. In the mouse genome, the hox11 family consists of at least three related genes, each of which possesses a highly conserved homeobox. To assist in elucidating the roles of this gene, a homologue was studied from Drosophila melanogaster. This gene, 311, shares similar identity to all three murine family members and contains the threonine residue in helix 3 of the homeodomain characteristic of the Hox11 family. It maps to a cluster of NK-homeobox genes which function in muscle development. Gene 311 exhibits a similar temporal pattern of expression to the NK genes in this cluster. Therefore, 311 may constitute part of a homeobox cluster in which the genes are both co-ordinately regulated and functionally related.
Assuntos
Drosophila melanogaster/genética , Genes Homeobox , Genes de Insetos , Sequência de Aminoácidos , Animais , Northern Blotting , Mapeamento Cromossômico , Drosophila melanogaster/crescimento & desenvolvimento , Humanos , Leucemia-Linfoma de Células T do Adulto/genética , Camundongos , Dados de Sequência Molecular , Família Multigênica , RNA Mensageiro/análise , Alinhamento de Sequência , Homologia de Sequência de AminoácidosRESUMO
A translocation involving human chromosome 10, band q24, in a subset of T-cell acute leukemias disrupts a region surrounding the putative oncogene HOX11, which encodes a protein with a homeodomain. The HOX11 protein binds to a specific DNA sequence, it localizes to the cell nucleus, and it transactivates transcription of a reporter gene linked to a cis-regulatory element, suggesting that HOX11 functions in vivo as a positive transcription activator. PCR analysis shows that the HOX11 homeodomain is a member of a distinct class of homeodomains, representatives of which occur in murine and Drosophila genomes. These all contain a threonine residue in place of the more common isoleucine or valine in helix 3 of the homeodomain. HOX11 therefore appears to belong to a family of DNA-binding transactivators of transcription.
Assuntos
Proteínas de Ligação a DNA/genética , Genes Homeobox , Proteínas de Homeodomínio , Proteínas Nucleares/genética , Proteínas Oncogênicas/genética , Fatores de Transcrição/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Sítios de Ligação , Núcleo Celular/metabolismo , Sequência Consenso , Genes de Insetos , Humanos , Camundongos , Dados de Sequência Molecular , Família Multigênica , Mutagênese Sítio-Dirigida , Oligodesoxirribonucleotídeos/química , Proteínas Proto-Oncogênicas , Proteínas Recombinantes/metabolismo , Relação Estrutura-Atividade , Ativação Transcricional , TransfecçãoRESUMO
Odorant detection is specifically mediated via receptor neurons in the olfactory mucosa but is a complex process involving a number of different cell types producing proteins of differing function. We have used the technique of subtractive hybridization cDNA cloning to identify novel genes expressed exclusively in the olfactory mucosa which may play a role in olfaction. Ten distinct groups of cDNA clones were identified which corresponded to mRNA transcripts highly expressed in rat olfactory mucosa but undetectable in thymus, kidney, lung, brain, spleen and liver. Some of these clones identify substructures in the mucosal tissue for which no other probes are currently available. Others identify novel mRNA species in the Bowman's glands. The predicted proteins for three of these clones are homologous to proteins which bind to either lipopolysaccharides (RYA3 and RY2G5) or to polychlorinated biphenyls (RYD5). In addition, while RYA3 and RY2G5 are highly homologous, they appear to be expressed in different parts of the mucosal tissue. The sequence homologies and subanatomical location of expression suggest that these proteins might interact with odorants before or after specific recognition by odorant receptors. Therefore, the olfactory mucosa may possess diverse, functionally-distinct odorant-binding proteins which recognize and bind separate classes of odorants.
Assuntos
Proteínas de Transporte/metabolismo , Mucosa Nasal/metabolismo , Receptores Odorantes , Células Receptoras Sensoriais/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas de Transporte/genética , DNA/genética , Ligantes , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , RNA Mensageiro/análise , Ratos , Ratos Endogâmicos F344 , Alinhamento de Sequência , Homologia de Sequência do Ácido NucleicoRESUMO
Olfactory reception occurs via the interaction of odorants with the chemosensory cilia of the olfactory receptor cells located in the nasal epithelium. The cDNA clones from mRNA specific to olfactory mucosa were studied. One of these clones, OBPII, encodes a secretory protein with significant homology to odorant-binding protein (OBP), a protein with broad odorant-binding ability, and is expressed in the lateral nasal gland, which is the site of expression of OBP. The OBPII sequence also shows significant homology to the VEG protein, which is thought to be involved in taste transduction. OBPII is a new member of the lipophilic molecule carrier protein family. The second cDNA clone encodes a novel homologue of glutathione peroxidase, an enzyme involved in cellular biotransformation pathways. Its expression appears to be localized to the Bowman's glands, the site of several previously identified olfactory-specific biotransformation enzymes.
Assuntos
Proteínas de Transporte/genética , Receptores Odorantes , Células Receptoras Sensoriais/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Northern Blotting , Southern Blotting , Proteínas de Transporte/metabolismo , Clonagem Molecular , DNA/genética , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos F344 , Alinhamento de SequênciaRESUMO
It has been previously demonstrated that the WDNM1 gene is downregulated in metastatic rat mammary adenocarcinomas. Detailed analysis of the structure of WDNM1 mRNA reveals the presence of two transcripts generated by alternative splicing of a single exon. The unspliced version possesses an in-frame termination codon which encodes a predicted protein of 18 amino acids. The spliced version encodes a predicted protein of 60 amino acids which exhibits strong homology to a family of proteins possessing a conserved arrangement of cysteine residues. This family includes several proteinase inhibitors suggesting that WDNM1 could encode a product with proteinase inhibiting capacity. Possible modulation in the level of this protein product could be related to overall proteinase activity of specific tumour cells which, in turn, could influence the invasive and metastatic potential of such cell populations.