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M Chaudhuri

Publications and source records attributed to M Chaudhuri.

At least 19 recordsLinked to original sources
PubMed1995

The phosphotyrosine interaction domain of SHC recognizes tyrosine-phosphorylated NPXY motif.

Reversible assembly of intracellular signaling complexes is, in some cases, mediated by direct binding of a Src homology 2 (SH2) domain of one protein to a phosphotyrosine moiety of another protein (Cantley, L. C., Auger, K. R., Carpenter, C. L., Duckworth, B., Graziani, A., Kapeller, R., and Soltoff, S. (1991) Cell 64, 281-302). Using a degenerate phosphotyrosine-containing peptide library, we showed that individual SH2 domains recognize phosphotyrosine in a specific sequence context to provide fidelity in signaling (Songyang, Z., Shoelson, S. E., Chaudhuri, M., Gish, G., Pawson, T., Haser, W. G., King, F., Roberts, T., Ratnofsky, S., Lechleider, R. J., Neel, B. G., Birge, R. B., Fajardo, J. E., Chou, M. M., Hanafusa, H., Schaffhausen, B., and Cantley, L. C. (1993) Cell 72, 767-778). Recently a second type of phosphotyrosine interaction domain (PID) or phosphotyrosine-binding domain (PTB) was discovered in the amino terminus of the SHC proto-oncoprotein (Kavanaugh, W. M., and Williams, L. (1994) Science 266, 1862-1865; Blaikie, P., Immanuel, D., Wu, J., Li, N., Yajnik, V., and Margolis, B. (1994) J. Biol. Chem. 269, 32031-32034). Here we demonstrate, using a phosphotyrosine peptide library, that the SHC PID domain preferentially binds to the sequence Asn-Pro-Xaa-phosphotyrosine. This motif is in agreement with sequences at sites implicated in in vivo SHC binding. These results indicate that while SH2 domains predominantly interact with specific residues carboxyl-terminal of phosphotyrosine, the PID domain has high specificity for residues amino-terminal of phosphotyrosine.

Amino Acid Sequence↗
PubMed1993

Phosphatidylinositol 3-kinase p85 SH2 domain specificity defined by direct phosphopeptide/SH2 domain binding.

We have developed a competition binding assay to quantify relative affinities of isolated Src-homology 2 (SH2) domains for phosphopeptide sequences. Eleven synthetic 11-12-amino acid phosphopeptides containing YMXM or YVXM recognition motifs bound to a PI 3-kinase p85 SH2 domain with highest affinities, including sequences surrounding phosphorylated tyrosines of the PDGF, CSF-1/c-Fms, and kit-encoded receptors, IRS-1, and polyoma middle T antigens; matched, unphosphorylated sequences did not bind. A scrambled YMXM phosphopeptide or sequences corresponding to the GAP or PLC-gamma SH2 domain binding motifs of the PDGF, FGF, and EGF receptors bound to the p85 SH2 domain with 30-100-fold reduced affinity, indicating that this affinity range confers specificity. Binding specificity was appropriately reversed with an SH2 domain from PLC-gamma: a phosphopeptide corresponding to the site surrounding PDGF receptor Tyr1021 binds with approximately 40-fold higher affinity than a YMXM-phosphopeptide. We conclude that essential features of specific phosphoprotein/SH2 domain interactions can be reconstituted using truncated versions of both the phosphoprotein (a phosphopeptide) and cognate SH2 domain-containing protein (the SH2 domain). SH2 domain binding specificity results from differences in affinity conferred by the linear sequence surrounding phosphotyrosine.

Amino Acid Sequence↗
PubMed1993

SH2 domains recognize specific phosphopeptide sequences.

A phosphopeptide library was used to determine the sequence specificity of the peptide-binding sites of SH2 domains. One group of SH2 domains (Src, Fyn, Lck, Fgr, Abl, Crk, and Nck) preferred sequences with the general motif pTyr-hydrophilic-hydrophilic-Ile/Pro while another group (SH2 domains of p85, phospholipase C-gamma, and SHPTP2) selected the general motif pTyr-hydrophobic-X-hydrophobic. Individual members of these groups selected unique sequences, except the Src subfamily (Src, Fyn, Lck, and Fgr), which all selected the sequence pTyr-Glu-Glu-Ile. The variability in SH2 domain sequences at likely sites of contact provides a structural basis for the phosphopeptide selectivity of these families. Possible in vivo binding sites of the SH2 domains are discussed.

Amino Acid Sequence↗
PubMed1993

SH2 domains exhibit high-affinity binding to tyrosine-phosphorylated peptides yet also exhibit rapid dissociation and exchange.

src homology 2 (SH2) domains of intracellular signaling molecules such as phospholipase C-gamma and phosphatidylinositol 3'-kinase-associated protein p85 represent recognition motifs for specific phosphotyrosine-containing regions on activated growth factor receptors. The binding of SH2 domains to activated growth factor receptors controls the interaction with signaling molecules and the regulation of their activities. In this report, we describe the kinetic parameters and binding affinities of SH2 domains of p85 toward short phosphotyrosine-containing peptides with the amino acid sequence motif YMXM, derived from a major insulin receptor substrate, IRS-1, by using real time biospecific interaction analysis (BIAcore). Associations were specific and of very high affinity, with dissociation constants of 0.3 to 3 nM, between phosphopeptides and the two separate SH2 domains contained within p85. Nonphosphorylated peptides showed no measurable binding, and the interactions were specific for the primary sequence very close to the phosphotyrosine residue. Moreover, the interactions between phosphopeptides and SH2 domains of other signaling molecules were of much lower affinity. Interestingly, the binding of the SH2 domains to the tyrosine-phosphorylated peptides was of high affinity as a result of a very high on rate, of 3 x 10(7) to 40 x 10(7)/M/s; at the same time, the rate of dissociation, of 0.11 to 0.19/s, was rapid, allowing for rapid exchange of associating proteins with the tyrosine phosphorylation sites.

Amino Acid Sequence↗
PubMed1992

YMXM motifs of IRS-1 define substrate specificity of the insulin receptor kinase.

Of 34 tyrosine residues in insulin receptor substrate 1 (IRS-1), 14 are adjacent to acidic residues, suggesting that they might be phosphorylation sites. Synthetic peptides corresponding to sequences surrounding these tyrosines were used as substrates of the insulin receptor kinase. Surprisingly six of these, each within YMXM motifs, were phosphorylated with greatest efficiency (Km, 24-92 microM; kcat/Km, 0.6-2.1 x 10(4) M-1.sec-1). Substituted YMXM peptides revealed a strong preference of the insulin receptor kinase for methionine at Y + 1 and Y + 3 positions. When phosphorylated, related YMXM sequences are recognition motifs for binding to proteins with src-homology (SH2) domains. The combined hydrophobic and flexible nature of methionine side chains adjacent to the targeted tyrosines provides a versatile contact for recognition by diverse proteins involved in signal transduction.

Amino Acid Sequence↗
PubMed1990

Structures of the N-linked oligosaccharides of Gp63, the major surface glycoprotein, from Leishmania mexicana amazonensis.

The extent of protein N-glycosylation in Leishmania mexicana amazonensis has been proposed to be a factor in the virulence of the parasite. The N-linked oligosaccharides of gp63, the major surface glycoprotein of L. mexicana amazonensis, were characterized after their release by hydrazinolysis, re-N-acetylation, and reduction with NaB3H4. High voltage paper electrophoresis of the reduced oligosaccharides revealed only neutral species. Gel-permeation chromatography on Bio-Gel P-4 yielded four fractions, and the oligosaccharides present were structurally characterized by sequential exoglycosidase digestion, fragmentation by acetolysis, and methylation analysis. Four major structures were found and were biantennary oligomannose type with compositions of Glc1Man6GlcNAc2 (La), Man6GlcNAc2 (Lb), Man5GlcNAc2 (Lc), and Man4GlcNAc2 (Ld). The largest oligosaccharide (La) was shown to contain a terminal glucopyranosyl residue on the alpha (1----3) arm. The biantennary oligomannose structures (Lb and Lc) and the glucosylated structure Glc1Man6GlcNAc2 (La) have not previously been reported as a component of a mature glycoprotein from any source.

Animals↗
PubMed1990

Urinary androgen concentrations and social status in two species of free ranging zebra (Equus burchelli and E. grevyi).

In both species of zebra, breeding males had higher urinary androgen concentrations (ng androgens/mg Cr) than did non-breeding bachelor males (30.0 +/- 5.0 (N = 9) versus 11.4 +/- 2.8, (N = 7) in the plains zebra; 19.0 +/- 2.2 (N = 17) versus 10.7 +/- 1.2 (N = 14) in the Grevy's zebra). In the more stable family structure of the plains zebra (single male non-territorial groups) variations in androgen concentrations could not be ascribed to any measured variable. In the Grevy's zebra, androgen values were significantly lower in samples taken from territorial (breeding) males which had temporarily abandoned their territories (N = 4) and the urinary androgen concentration for a male on his territory was negatively correlated with the time since females last visited the territory.

Androgens↗
PubMed1989

Surface acid proteinase (gp63) of Leishmania mexicana. A metalloenzyme capable of protecting liposome-encapsulated proteins from phagolysosomal degradation by macrophages.

Acid proteinase activity is associated with the major surface glycoprotein (gp63) of both extracellular promastigotes and intracellular amastigotes of the parasitic protozoan, Leishmania mexicana. The enzyme purified by monoclonal affinity chromatography from promastigotes is strongly inhibited by metal ion chelators, which is reversible by the addition of Zn(II). This proteinase loses its activity after dialysis against 1,10-phenanthroline. The apoenzyme thus prepared is reactivated substantially by Zn(II) and partially by Cu(II), Cd(II), Co(II), or Ni(II). From the recently published structure of the gene encoding gp63, we identify hitherto unrecognized sequences, which can be aligned to the consensus zinc-binding sites of other known metalloproteinases. Anti-gp63 polyclonal antibodies, but not the monoclonals, precipitate similar molecules from amastigotes. These molecules differ slightly from gp63 in electrophoretic mobility but have similar endopeptidase activity. Phagolysosomal degradation by macrophages of proteins entrapped in liposomes is prevented by coating them with native gp63. This protection is lost with heat denaturation of gp63 to kill its enzymatic activity. The proteolytic activity of the metalloenzyme on the surface of these parasites may thus protect their membrane from cytolytic damages during their survival, differentiation, and multiplication in the phagolysosomes of macrophages.

Animals↗
PubMed1989

Factor interactions with the simian virus 40 early pre-mRNA influence branch site selection and alternative splicing.

To study the interaction of splicing factors with the simian virus 40 early-region pre-RNA, which can be alternatively spliced to produce large T and small t mRNAs, we used an in vitro RNase protection assay that defines the 5' boundaries of factor-RNA interactions. Protection products reflecting factor interactions with the large T and small t 5' splice sites and with the multiple lariat branch site region were characterized. All protection products were detected very early in the splicing reaction, before the appearance of spliced RNAs. However, protection of the large T 5' splice site was detected well before small t 5' splice site and branch site protection products, which appeared simultaneously. Oligonucleotide-targeted degradation of small nuclear RNAs (snRNAs) revealed that protection of the branch site region, which occurred at multiple sites, required intact U2 snRNA and was enhanced by U1 snRNA, while protection of the large T and small t 5' splice sites required both U1 and U2 snRNAs. Analysis of several pre-RNAs containing mutations in the branch site region suggests that factor interactions involving the multiple copies of the branch site consensus determine the selection of branch points, which is an important factor in the selection of alternative splicing pathways.

Binding Sites↗
PubMed1988

Antagonists of luteinizing hormone releasing hormone bind to rat mast cells and induce histamine release.

It was reported previously that administration of certain synthetic antagonists of LHRH to rats produced allergy-like symptoms that were attributed to their histamine releasing action. In the present study the interaction of LHRH analogs with rat peritoneal mast cells was investigated in vitro. Potent antagonists of LHRH showed strong in vitro histamine releasing activity from rat peritoneal mast cells. Membrane preparations of rat pituitary glands showed specific binding of radioiodinated LHRH antagonist as well as LHRH agonist. However, rat peritoneal mast cells and membrane preparations from those cells bound antagonist but not the agonist. Furthermore, the LHRH antagonist did not bind to membranes prepared from tissues such as prostate, liver, kidney, and brain. Competitive displacement curves of the [125I]-antagonist with different LHRH analogs showed that the ability of the analogs to compete for binding sites on mast cells was related to their histamine releasing activity. We conclude that histamine release from rat mast cells induced by LHRH analogs is mediated by specific binding of the active peptides to cell membranes. Furthermore, using rat mast cells, the binding assay in conjunction with histamine releasing assay may be utilized to predict the in vivo histamine releasing potential of new LHRH peptides which are of clinical importance.

Animals↗
PubMed1988

Induced type-B reticulum cell neoplasia in mice III. The importance of T-cell proliferation and cellular relocation in accessory cell transformation.

After the transfer of spleen cells from old CBA/T6T6 mice (greater than 75 weeks) into young syngeneic CBA/Ca recipients there usually follows a selective expansion of the donor T-cell population and the emergence of type B reticulum cell neoplasms (RCN-B), also of donor origin though probably derived not from the T-cells but from lymphoid dendritic accessory cells. As few as one million injected cells led to significant donor T-cell hyperplasia and tumour induction. Injection of cells from young donors did not have such consequences. Similar tumours were induced by transferring syngeneic cells in both C57BL and DBA/2 mice, although in the latter strain there was no requirement for the injected cells to derive from old donors. It appeared that T-cell proliferation was independent of donor accessory cells or RCN-B induction, since injection of enriched T-cells led to few tumours, although the T-cell chimaerism was indistinguishable from that in recipients of unseparated spleen cells. Development of tumours, however, seemed to be dependent upon stimulated T-cells. Recipients of spleen cells from old T-cell-deprived mice did not develop tumours; conversely, tumours, mostly of donor origin, were induced in recipients of young syngeneic cells when an extrinsic stimulus to T-cell proliferation was provided by continued allostimulation. The apparent selectivity of tumorigenesis for donor cells has led to the proposal that cellular relocation, as a result of transfer, may be an important predisposing factor in malignant transformation in circumstances of T-cell stimulation provided by antigenic challenge or by transfer of T-cells from old donors.

Age Factors↗
PubMed1988

Urinary steroid concentrations during natural and gonadotrophin-induced oestrus and pregnancy in the giant panda (Ailuropoda melanoleuca).

Urinary concentrations of conjugated oestrone and pregnanediol-3-glucuronide were measured during and after spontaneous and induced oestrus and during pregnancy. Behavioural oestrus was preceded by a rise in oestrone values from less than 10 ng/mg creatinine (Cr) to peaks of 45 ng/mg Cr. Maximal lordotic response and mating activity coincided with the decline in oestrone levels. After presumed ovulation, urinary pregnanediol glucuronide concentrations increased from less than 5 to 15-30 ng/mg Cr. Further increases in this steroid (to 60-80 ng/mg Cr) occurred 114 days after mating, presumably coincident with implantation. These high levels of pregnanediol glucuronide were maintained for 3 weeks, began to decline 1 week before parturition and fell to a nadir (less than 5 ng/mg Cr) immediately after delivery. When FSH was administered i.m. for 5 days, urinary oestrone values rose markedly and were maximal (580 ng/mg Cr) on Day 7. Mating first occurred on Day 20 and 500 i.u. hCG were given i.m. Urinary pregnanediol glucuronide levels during the next 5 months were similar to those in the previous year during pregnancy with values rising 105-108 days after mating. However, no birth occurred. These results support the suggestion that pandas exhibit delayed implantation and demonstrate that the panda is responsive to exogenous gonadotrophins.

Animals↗
PubMed1986

The capability for regulation of insulin secretion by somatostatin in purified pancreatic islet B cells during aging.

Pancreatic islet B cells from Sprague-Dawley and Fisher 344 rats aged 3-27 months were separated from A and D cells by centrifugation over a linear percoll density gradient, and incubated in vitro with various concentrations of glucose and somatostatin. Elevation of glucose concentration in the incubation medium from 2.6 to 16.7 mM provokes an insulin secretory response that is independent of rat donor age. Inhibition of the insulin secretory response by somatostatin is independent of rat donor age beyond 12 months. These data indicate that the impaired regulation of insulin secretion during aging observed previously in vivo and in vitro in intact islets may not be intrinsic to the B cells, but instead reflect changes in islet paracrine regulatory mechanisms that relate to the quality and/or quantity of endogenous somatostatin and/or glucagon.

Aging↗
PubMed1986

Regulation of insulin secretion by glucose during aging.

The purpose of this study was to investigate the sensitivity of insulin secretion to glucose and its possible relationship to changes in islet glucose utilization during aging. Pancreatic islets of Langerhans were isolated from male Sprague-Dawley rats aged 2- to 24-months and perifused in vitro with glucose. Insulin concentration was determined by radioimmunoassay. Glucose utilization was assessed by measuring the rate of conversion of 3H-5-D-glucose to 3H-H20. The sensitivity of isolated, perifused islets to glucose was reduced during aging. Less glucose metabolism, however, may be required to elicit comparable release of insulin by islets from old than from young rats.

Aging↗
PubMed1985

Pygopagus twins.

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Adult↗
PubMed1984

Neoplasms arising in CBA mice after transfer of spleen cells from syngeneic old donors.

Young (15-week-old) CBA/Ca mice were injected intravenously with spleen cells from individual young (15-week-old) or old (18-24-month-old) CBA/T6T6 mice. Samples of peripheral blood were taken at monthly intervals and cultured with phytohaemagglutinin (PHA) to stimulate T lymphocytes into mitosis. In the recipients of young lymphocytes, the percentage of donor cells dividing in these cultures remained low throughout the experiment, but in the recipients of old spleen cells, after an initial period when the percentage of donor cells declined, there was a marked increase in the percentage of donor cells. The interval between the injection and the increase in the proportion of donor cells was very variable. Ten of the 12 recipients of old lymphocytes developed tumours involving the spleen and mesenteric lymph node. They resembled type B reticulum cell neoplasms as described by Dunn & Deringer (1968), and all but two were transplantable. In addition, one mouse that had no evidence of tumour on histological examination nevertheless gave rise to a transplantable tumour. The five tumours on which chromosomal analysis was carried out proved to be of old donor cell origin. Two out of the five recipients of young cells also eventually developed tumours, but these arose later than the others, had a more granulocytic character, and did not transplant.

Age Factors↗
PubMed1983

Recirculating and sessile B cell populations in normal and CBA/N mice.

Chromosomally distinguishable syngeneic mice were parabiosed and the resultant chimerism was followed for 6 weeks in the lymphoid organs, by culturing their cells with polyclonal mitogens, lipopolysaccharide (LPS) for B cells and phytohemagglutinin (PHA) for T cells. As expected of a recirculating population, the T cells equilibrated completely. The B cells in lymph nodes (LN) and Peyer's patches (PP) also equilibrated completely, suggesting that they too are recirculating. B cells in the spleen and blood, however, did not equilibrate over this period. After separation of parabiosed mice, the percentage of partner cells in both the recirculating T and B lymphocyte populations declined steadily, but it continued to rise in the LPS-responsive populations in spleen and peripheral blood suggesting that they were derived from precursor populations which were themselves chimeric. Injection of lymphocytes into CBA/Ca or CBA/N mice showed that LPS-responsive populations in LN and spleen localized differently. These results have been interpreted as demonstrating two major populations of LPS-responsive B lymphocytes in the mouse, one recirculating and the other sessile. The recirculating population appears to be the only LPS-responsive population in LN and PP. In the spleen, however, the recirculating cells constitute about a quarter of the LPS-responsive cells, while the rest are sessile cells. The relationship between these two populations has yet to be clarified. CBA/N mice are deficient in both populations but the sessile one appears to be more severely depleted.

Animals↗