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P de Santa Barbara

Publications and source records attributed to P de Santa Barbara.

10 recordsLinked to original sources
PubMed2003

Development and differentiation of the intestinal epithelium.

The gastrointestinal tract develops from a simple tube to a complex organ with patterns of differentiation along four axes of asymmetry. The organ is composed of all three germ layers signaling to each other during development to form the adult structure. The gut epithelium is a constitutively developing tissue, constantly differentiating from a stem cell in a progenitor pool throughout the life of the organism. Signals from the adjacent mesoderm and between epithelial cells are required for normal orderly development/differentiation, homeostasis, and apoptosis. Embryonically important patterning factors are used during adult stages for these processes. Such critical pathways as the hedgehog, bone morphogenetic protein, Notch, Sox, and Wnt systems are used both in embryologic and adult times of gut development. We focus on and review the roles of these factors in gut epithelial cell development and differentiation.

Adult↗
PubMed2001

Steroidogenic factor-1 contributes to the cyclic-adenosine monophosphate down-regulation of human SRY gene expression.

In mammals, male sex determination is initiated by SRY (sex-determining region of the Y chromosome) gene expression and followed by testicular development. This study describes specific down-regulation of the human SRY gene transcription by cAMP stimulation using reverse transcription-polymerase chain reaction experiments. Using transfection experiments, conserved nuclear hormone receptor (NHR1) and Sp1 consensus binding sites were identified as essential for this cAMP transcriptional response. Steroidogenic factor-1 (SF-1), a component of the sex-determination cascade, binds specifically to the NHR1 site and activates the SRY promoter. Activation of SF-1 was abolished by cAMP pretreatment of the cells, suggesting a possible effect of cAMP on the SF-1 protein itself. Indeed, human SF-1 protein contains at least two in vitro cAMP-dependent protein kinase (PKA) phosphorylation sites, leading after phosphorylation to a modification of both DNA-binding activity and interaction with general transcription factors such as Sp1. Taken together, these data suggest that cAMP responsiveness of human SRY promoter involves both SF-1 and Sp1 sites and could act via PKA phosphorylation of the SF-1 protein itself.

Cyclic AMP↗
PubMed2000

Expression and subcellular localization of SF-1, SOX9, WT1, and AMH proteins during early human testicular development.

Many transcription factors have been identified and implicated in male sex determination pathway. Specifically involved in Sertoli cell differentiation and subsequent anti-Müllerian hormone (AMH) secretion in eutherian mammals, they include steroidogenic factor-1 (SF-1), SOX9 (SRY HMG box related gene 9), WT1 (Wilms' tumor 1), and GATA-4 (a zinc finger transcription factor). These factors have been described to execute their function in the male sex determination pathway by controlling AMH transcriptional expression. To understand the hierarchies of these factors and their involvement in the developing testis, for the first time we show the expression and subcellular localization of these factors by immunohistochemistry in the early human testis during embryogenesis compared with AMH expression. If these studies do not refute their possible synergistic interaction to control AMH expression in human embryo, they also reveal a new sexual dimorphism in SOX9 expression during the sex determination process. We show that SOX9 sex specifically shifts from the cytoplasmic to the nuclear compartment at the time of testis differentiation and AMH expression. Putative models for this subcellular distribution are discussed.

Anti-Mullerian Hormone↗
PubMed1999

[Sex determination in mammals: state of the art].

Sex determination relies on the translation of chromosomal sex established at fertilisation into gonadal sex (testis or ovary), and later into somatic sex (male or female) under the control of gonadal hormone secretions. The aim of the current review will be to highlight our knowledge of the key events which, in the presence of a Y chromosome, induce the organisation of the developing epithelial cells located inside the genital ridges into testicular cords. Many groups have tried to define the molecules relevant to this process, with a double goal: unravelling a molecular pathway which leads to cell fate decision (Sertoli cell in this particular case) during development; improving the establishment of a diagnosis and subsequent medical management in cases where chromosomal, gonadal and then somatic sexes are discordant. Recent progress made in this area will be depicted, with the introduction of several pieces to this developmental jigsaw puzzle.

Animals↗
PubMed1998

Steroidogenic factor-1 regulates transcription of the human anti-müllerian hormone receptor.

Anti-müllerian hormone type II receptor (AMHRII) is a serine/threonine receptor and a member of type II receptors of the transforming growth factor beta superfamily. AMHRII has been recently identified in humans, mice, rats, and rabbits. In the male embryo, the AMHRII gene has been shown to be expressed in Sertoli's cells, in Leydig's cells and in the mesenchymal cells surrounding the müllerian duct. To determine the functional region of the AMHRII promoter as well as the factors controlling AMHRII gene expression, we used a 1.1-kilobase DNA fragment from the 5'-flanking region of the human AMHRII gene to generate a series of deletion or mutation and analyzed the resulting transcriptional activities after transfection of the NT2/D1 teratocarcinoma cell line. Our results indicate that maximal expression of the AMHRII promoter in this particular cell line, a cell line positive for endogenous AMHRII expression, requires a conserved estrogen receptor half-site element (AGGTCA) identical to the binding element for steroidogenic factor-1 (SF-1). Studies of this SF-1 binding element using gel mobility shift, antibody supershift assays, and transient transfections of reporter constructs indicate that SF-1 can bind and transactivate the AMHRII promoter. Finally, SF-1 protein expression in human male embryos was shown to display a good coincidence with the previously reported AMHRII expression profile. We then propose that SF-1 may be a key transcriptional regulator of AMHRII gene expression during early human development.

Animals↗
PubMed1998

Characterization of two Sp1 binding sites of the human sex determining SRY promoter.

To investigate the molecular basis of the human SRY gene regulation, we have examined the significance of two potential binding sites for the transcription factor Sp1 (Sp1A: -124 to -131 and Sp1B: -147 to -154) by DNase I footprinting and gel mobility shift assays. Cotransfection experiments in Drosophila SL2 cells implicated Sp1 protein in the transcriptional activation of the SRY promoter.

Animals↗
PubMed1998

Phosphorylation of an N-terminal motif enhances DNA-binding activity of the human SRY protein.

Of the several strategies that eukaryotes have evolved to modulate transcription factor activity, phosphorylation is regarded as one of the major mechanisms in signal-dependent transcriptional control. To conclusively demonstrate that the human sex-determining gene SRY is affected by such a post-translational control mechanism, we have analyzed its phosphorylation status in living cells. In the present study, we show that the cyclic AMP-dependent protein kinase (PKA) phosphorylates the human SRY protein in vitro as well as in vivo on serine residues located in the N-terminal part of the protein. This phosphorylation event was shown to positively regulate SRY DNA-binding activity and to enhance the ability of SRY to inhibit a basal promoter activity located downstream of an SRY DNA-binding site concatamer. Together these results strongly support the hypothesis that human SRY is a natural substrate for PKA in vivo and that this phosphorylation significantly modulates its major activity, DNA-binding, thereby possibly altering its biological function.

Amino Acid Sequence↗
PubMed1997

The human testis determining factor SRY binds a nuclear factor containing PDZ protein interaction domains.

The human Y-linked testis determining gene SRY encodes a protein with a DNA binding domain from the high mobility group box family. To date, no function has been assigned to amino acid sequences located outside this DNA binding motif. Here, we identify in a yeast two-hybrid screen a PDZ protein termed SIP-1, as an interacting protein with human SRY. In vitro, biochemical analysis, immunoprecipitation experiments, as well as expression of SIP-1 in human embryonic testis confirm that the two proteins can interact together. Interacting domains were mapped to the C-terminal seven amino acids of SRY and to the PDZ domains of SIP-1, respectively. We hypothesize that SIP-1 could connect SRY to other transcription factors providing SRY for its missing trans-regulation domain.

Amino Acid Sequence↗
PubMed1996

Change in the N-terminal domain conformation of annexin I that correlates with liposome aggregation is impaired by Ser-27 to Glu mutation that mimics phosphorylation.

Annexin I is a member of the annexin family of calcium-dependent membrane binding proteins. The core domain of these proteins is similar in all annexins but the N-terminal domain is specific for each member. This domain is thought to regulate annexin function through phosphorylation. In annexin I, Ser-27 is one of the amino acids that can be phosphorylated by protein kinase C. Phosphorylations are thought to regulate some annexin I functions by increasing calcium requirement. Two mutants were prepared in this study: S27E and S27A proteins. The first mimics phosphorylation while the second prevents phosphorylation at residue 27. Wild-type annexin I and S27A mutant protein showed the same calcium dependence for phospholipid vesicles aggregation, while S27E mutant protein showed a higher calcium requirement and a low maximal extent of aggregation. By contrast, liposome binding and self-association required identical calcium concentrations for the wild-type and the two mutant proteins. To examine whether the regulation observed is due to modification of the N-terminal structure, we investigated conformational changes by using two approaches. Firstly we analysed proteinase sensibility. Limited proteolysis of the N-terminal tail showed similar patterns for the three proteins. Using drastic conditions of proteolysis, we observed strong resistance of the core domain to digestion in the presence of calcium. Secondly, since Ser-27 is located on the N-terminal domain that contains a tryptophan located at position 12, the fluorescence of this residue was analysed during Ca2+-binding of wild-type annexin I and S27E mutant protein. The results demonstrated that Ca2+ induces a slight change in the Trp environment of wild-type annexin I, corresponding to a burying of the residue. No changes in fluorescence features were observed with S27E mutant protein. The results obtained show that phosphorylation of the N-terminal tail plays a regulatory role in phospholipid vesicle aggregation, which is based on a mechanism distinct from protein self-association. This phosphorylation induces a conformational change in the tail probably related to aggregation property.

Annexin A1↗