RESUMO
Major developmental transitions in thymocyte differentiation are accompanied by sharp alterations in cAMP metabolism. We have analyzed the cAMP accumulation responses of cell populations representing successive stages of T-cell development, namely: immature TcR- thymocytes from SCID mice, proliferating cortical blasts, small cortical thymocytes, medullary thymocytes and peripheral T cells. We find that all classes of thymocytes exhibit higher cAMP synthesis in response to forskolin than peripheral T cells. In immature TcR- thymocytes, this high capacity is buffered by efficient phosphodiesterase activity, but in CD4+CD8+TcRlow thymocytes, phosphodiesterase activity becomes much less effective. Phosphodiesterase activity then rises again after positive selection. The ability of thymocytes to respond to prostaglandin E is regulated distinctly from their ability to respond to forskolin. Unlike forskolin, PGE1 induces cAMP synthesis to similar levels in all classes of thymocytes, possibly due to partial activation of phosphodiesterase in cortical thymocytes by PGE1. Finally, we report a novel effect of Ca2+/protein kinase C signaling on cAMP accumulation, which occurs selectively in the proliferating cortical blasts.
Assuntos
AMP Cíclico/fisiologia , Transdução de Sinais , Linfócitos T/fisiologia , Timo/fisiologia , Adenilil Ciclases/metabolismo , Alprostadil/farmacologia , Animais , Cálcio/metabolismo , Células Cultivadas , Colforsina/farmacologia , Citometria de Fluxo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos SCID , Índice Mitótico , Proteína Quinase C/metabolismo , Subpopulações de Linfócitos T/efeitos dos fármacos , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/fisiologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Acetato de Tetradecanoilforbol/farmacologia , Timo/imunologiaRESUMO
Two Drosophila melanogaster vitelline membrane protein-encoding genes (VM), located at polytene band positions 26A and 34C, have been cloned and comparatively characterized at the nucleotide level. Sequence analysis of genomic and cDNA clones for the two genes, VM26A.1 and VM34C.1, indicates that both are similarly organized with a central highly conserved domain [Scherer et al., Dev. Biol. 130 (1988) 786-788] which is flanked by unrelated regions, and that both genes lack introns. Comparison of the upstream regions reveals that both VM genes contain a hepatmeric element identical to one associated with the D. melanogaster yolk protein-encoding genes (YP). This heptamer occurs in the specific 5' flanking region responsible for ovarian temporal- and tissue-specific control in both VM and YP genes. A putative chorion transcription factor 2 site is also associated with an upstream control element of VM26A.1, but not with any sequenced portion of VM34C.1.
Assuntos
Proteínas de Drosophila , Drosophila melanogaster/genética , Proteínas do Ovo/genética , Proteínas de Membrana/genética , Membrana Vitelina , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , DNA Complementar , Dados de Sequência Molecular , Mapeamento por Restrição , Homologia de Sequência do Ácido NucleicoRESUMO
We have cloned full-length DDC cDNAs from a human hepatoma cDNA library [DDC; dopa decarboxylase; aromatic-L-amino acid decarboxylase, EC 4.1.1.28]. The protein encoded by hepatoma cells is the same as that encoded by adrenal chromaffin derived pheochromocytoma cells, despite reported differences in biochemical properties. We have confirmed the location of the DDC gene to chromosome 7 using a new panel of somatic cell hybrids, and we have localized the gene to band p11 on chromosome 7 by fluorescent in situ hybridization. The human gene retains 65% amino acid identity with Drosophila DDC (Accession No. X04426) and considerable structural similarity with other enzymes (F.R. Jackson, 1990, J. Mol. Evol. 31:325-329, and references therein).
Assuntos
Cromossomos Humanos Par 7 , DNA/genética , Dopa Descarboxilase/genética , Animais , Sequência de Bases , Southern Blotting , Mapeamento Cromossômico , Cricetinae , Cricetulus , Corantes Fluorescentes , Genes , Células HeLa , Humanos , Células Híbridas , Fígado/química , Dados de Sequência Molecular , Hibridização de Ácido NucleicoRESUMO
Two Drosophila vitelline membrane (VM) genes located at polytene band positions 26A and 34C have been characterized at the nucleotide level. Sequence comparison of the two genes VM26A.1 and VM34C.1 has revealed a similar 114 base pair region centrally located in the coding regions of both genes. The conserved region has a 91% homology at the nucleic acid level and a 100% conservation at the amino acid level. This suggests a common evolutionary origin for these VM genes and indicates that a strong selective pressure exists to maintain a specific polypeptide sequence in a domain of the proteins.
Assuntos
Drosophila melanogaster/genética , Membrana Vitelina/fisiologia , Animais , Sequência de Bases , Proteínas de Membrana/genética , Dados de Sequência Molecular , Sequências Repetitivas de Ácido NucleicoRESUMO
cDNA clones for two Drosophila vitelline membrane genes have been identified on the basis of: (i) stage and tissue specificity of transcription and (ii) size and amino acid content of the translation product. Cross-hybridization data suggest that DmcMM99 and DmcMM115 are members of a multi-gene family which includes at least three members, all of which reside on the left arm of the second chromosome. DmcMM99 and DmcMM115 originate from polytene band positions 34C and 26A, respectively. A third, cross-hybridizing gene resides at position 32EF. Southern analysis of a genomic clone, lambda LS1, homologous to DmcMM115, indicates that two vitelline membrane genes may be clustered at the 26A site.
