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Research Division, Joslin Diabetes Center (X.J.S., S.P., Y.Z.,
L.Y., D.B., M.G.M., E.G., M.F.W.) and Department of Medicine
Harvard Medical School Boston, Massachusetts 02215
Laboratory of Cell and Molecular Biology (L-M.Y., J.H.P.)
National Institutes of Health Bethesda, Maryland 20892
Mammalian Genetics Laboratory (N.G.C., N.A.J.) ABL-Basic
Research Program National Cancer Institute-Frederick Cancer
Research and Development Center Frederick, Maryland 21702
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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In a few cases, notably insulin and insulin-like growth factor 1
(IGF-1), receptor autophosphorylation correlates closely with increased
kinase activity but poorly with the recruitment of most SH2-proteins to
the receptor (6, 7). In contrast, tyrosine phosphorylation of the IRS
proteins (IRS-1 and IRS-2) provides an interface between these
receptors and various SH2-proteins (8, 9). IRS proteins are also
phosphorylated by Jak kinases, which are activated by the receptors for
various cytokines, including interleukins (IL-2, IL-4, 3IL-9,
IL-13), interferons (IFN
, IFNß and IFN
), and GH. During insulin
stimulation the phosphorylation of multiple tyrosine residues in the
COOH terminus of IRS-1 enables the binding of several SH2 proteins,
including the PI-3 kinase-regulatory subunits
(p85/p85ß/p55PIK), GRB-2, nck, c-fyn, and
SHPTP2 (10, 11, 12, 13, 14). As a consequence of these and other interactions,
IRS-1 mediates multiple downstream signals, including the direct
activation of PI-3 kinase and SHPTP2, the indirect stimulation of
mitogen-activated protein kinase and p70s6k, and other
events that regulate gene expression and stimulate protein synthesis,
mitogenesis, and glucose transport (6, 15, 16, 17, 18, 19, 20).
IRS-2 was difficult to identify using several standard approaches, including expression screening, RT-PCR, and low stringency cDNA or genomic screening. However, purification of 4PS, an insulin/IL-4 receptor substrate in FDC-P2 cells, allowed the cloning of a cDNA that encodes a protein with several structural and functional features in common with IRS-1 (21). IRS-2 appears to be especially important in mice lacking IRS-1, as IRS1(-/-) mice survive, reproduce, and display only mild insulin resistance (22, 23). Hepatocytes from these animals reveal increased tyrosine phosphorylation of IRS-2 and retain a significant responsiveness to insulin and IGF-1 (22, 23). However, muscle from the IRS1(-/-) mouse is significantly insulin resistant and does not display a compensatory increase in IRS-2 phosphorylation (24). Thus, the distinct expression of IRS-1 and IRS-2 may further contribute to their unique signaling potential and importance for survival.
In this paper we characterize and analyze the murine IRS-2 gene and investigate the potential for alternate signaling by IRS-1 and IRS-2 in different cell types and in response to different upstream signals. The IRS-2 gene is located on murine chromosome 8 in a location close to the insulin receptor; the coding region is contained in a single exon. Although IRS-2 possesses similar structural features to IRS-1, the tyrosine phosphorylation of IRS-1 and IRS-2 by insulin and IL-4 is qualitatively different.
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RESULTS |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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). Restriction mapping and PCR
analysis confirmed that the fragments obtained from the genomic DNA
corresponded exactly to the partial cDNA sequences obtained previously
from various murine libraries (Fig. 1A). Translation of the open
reading frame obtained from the genomic sequence revealed 1324 amino
acids, including three additional residues at the COOH terminus that
were not previously described (Fig. 1B) (21). The 5'-untranslated
region of the IRS-2 gene contains two GC-rich regions that proved
difficult to sequence accurately and remain undefined (Fig. 1B).
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). This pattern is
consistent with the conclusion that IRS-2 is encoded by a single gene
in the 129 mouse as diagramed in Fig. 1A
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), up to 181 mice were characterized for some pairs of
markers; each locus was analyzed in pairwise combinations for
recombination frequencies using the additional data. The ratios of the
total number of mice exhibiting recombinant chromosomes to the
total number of mice analyzed for each pair of loci suggests the
following gene order: centromere
Insr
(1/95)Irs2 (0/142)Col4al
(8/181)Plat (Fig. 2B). The lack of recombinations
detected between Irs2 and Col4al in this number
of animal suggests that the two loci are within 2.1 centiMorgans (cM)
of each other at the 95% confidence limit. The recombination
frequencies reported as the genetic distance in centiMorgans ±
SE is 1.1 ± 1.1 between the Insr loci and
Irs2/Col4al, and 4.4 ± 1.5 between
Irs2/Col4al and Plat (Fig. 2B).
Downstream Signaling Specificity of IRS-1 and IRS-2
The amino acid sequence identity in the COOH terminus of IRS-2 and
IRS-1 is only 35%, which arises largely from similar tyrosine
phosphorylation motifs surrounded by variable stretches of amino acid
sequence (21). To explore possible signaling diversity in IRS-1 and
IRS-2 at the level of tyrosine phosphorylation, we prepared a panel of
GST-fusion proteins containing SH2-domains from various signaling
molecules and tested their ability to bind to IRS-2 or IRS-1 expressed
in 32DIR/IL4R cells. After insulin or IL-4 stimulation,
cell lysates were incubated with GST-SH2 fusion proteins and the
associated IRS-2 and IRS-1 were measured by immunoblotting with PY
(25).
The NH2-terminal SH2 domain of p85 bound strongly to both
IRS-2 and IRS-1 during stimulation with insulin (Fig. 3); insulin and IL-4 stimulated similar amounts of IRS-2
binding to p85, while insulin stimulated more binding to IRS-1 than
IL-4 did. Similar results were observed with the SH2 domain of
c-fyn. However, the SH2 domains from Crk, phospholipase C,
and GRB-2 revealed functional differences. Each SH2 domain bound IRS-1
more strongly than IRS-2 during insulin and IL-4 stimulation. Moreover,
IRS-1 bound more tightly during insulin stimulation than during IL-4
stimulation, whereas IRS-2 bound equally during insulin and IL-4 (Fig. 3). The SH2 domains of Abl and SHP-2 associated only with IRS-1, and
then only during insulin stimulation (Fig. 3). These results suggest
that IRS-2 and IRS-1 may be phosphorylated differently by various
receptors and engage a unique cohort of SH2 proteins.
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PY during insulin stimulation (Fig. 4A). However, after differentiation, the amount of IRS-1
increased dramatically, and insulin robustly stimulated its tyrosine
phosphorylation; however, IRS-2 was barely detected and its
insulin-stimulated tyrosine phosphorylation was quite low (Fig. 4A).
Consistent with this observation, IRS-1 was the predominant
phosphorylated IRS protein in isolated rat adipocytes during insulin
stimulation; however, a small amount of p85 was detected in IRS2
immunoprecipitates after insulin stimulation, suggesting that a small
amount of IRS-2 is expressed in this background (Fig. 4B). Whether this
low expression of IRS-2 is sufficient to rescue insulin signaling in
murine adipocytes from the IRS-1(-/-) mouse is unknown,
but future experiments should reveal whether other pathways such as the
tyrosine phosphorylation of p60 or Gab1 are involved (27, 28).
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). This change in relative
expression continued until IRS-1 predominated in the fully
differentiated P19 neurons (6 days after retinoic acid removal).
Consistent with these results, IGF-1 strongly mediated IRS-1
phosphorylation in the P19 neurons and IRS-1 associated with p85 (Fig. 5). It will be interesting to determine whether the reduced expression
of IRS-2 is required for retinoic acid-induced differentiation in P19
cells.
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). Interestingly,
cross-linking surface IgM with IgM induced weak (relative to
insulin) tyrosine phosphorylation of IRS-2 (Fig. 6, lanes g-i). The
effect of IgM was specific as nonimmune serum had no effect,
suggesting that IRS-2 may function downstream of the B cell antigen
receptor (Fig. 6, lanes a-c). We also observed the tyrosine
phosphorylation of IRS-2 in isolated B cells during anti-IgM
stimulation (data not shown).
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). However, IRS-2 was conspicuously
absent from a few cell lines of lymphoid origin, including Daudi and
ABMC1 (B cell lines), and CT6 and EL4 (T cell lines). IRS-2 was barely
detected in rat testis and surprisingly weak in 3T3-L1 cells and rat
adipocytes (Table 1). By comparison, IRS-1 was also broadly expressed;
however, it was absent from several lymphoid cell lines and all cells
of the myeloid lineage that were tested (Table 1). Differential
expression was also observed in several carcinoma cell lines: IRS-2 was
highly expressed in three colon carcinoma cells (clone-A, CX-1, RKO),
whereas only clone A coexpressed IRS-1; both IRS-1 and IRS-2 were
expressed relatively highly in several mammary carcinoma cell lines
where they may contribute to their prolonged survival (Table 1).
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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IRS-1 and IRS-2 appear to interact with a similar cohort of cellular membrane receptors, including insulin/IGF-1, several ILs, IFNs, and GH (9). During stimulation of these receptors with their cognate ligands, the IRS proteins undergo tyrosine phosphorylation and associate with signaling proteins that contain SH2-proteins. Although the overall amino acid sequence identity between IRS-2 and IRS-1 is 43%, the alignment shows regions of striking homology within the NH2-terminal third of the molecule (21). Translation of the coding region of IRS-2 reveals at least three functional regions. Two highly conserved regions occur in the NH2-terminal portion of IRS-1 and IRS-2: the first IRS-Homology region (IH1PH) contains a PH domain; the second, IH2PTB, contains a PTB domain (32). Recent experiments indicate that both regions couple IRS-1 to the insulin receptor in 32D cells; however, the IH1PH region provides the most efficient coupling (33). Another region (between residues 313 and 462 of IRS-1) was aligned recently to the PTB domain in Shc and designated the "SAIN" domain (34, 35, 36); however, this region is poorly conserved between IRS-1 and IRS-2 and does not function as a PTB domain (33).
By contrast, the COOH-terminal regions of IRS-1 and IRS-2 are poorly conserved, displaying only 35% identity (21). The middle of IRS-2 contains a unique region that interacts specifically with the phosphorylated regulatory loop of the insulin receptor (35). This region, which is absent from IRS-1, may alter the phosphorylation pattern of the COOH terminus by restricting the flexibility of IRS-2 in the catalytic domain. On the other hand, several tyrosine phosphorylation sites in IRS-2 align with similarly spaced sites in IRS-1 (21). In several cases, the amino acid sequence around the tyrosine residues are nearly identical in IRS-2 and IRS-1, including the NH2-terminal acidic residues and the COOH-terminal hydrophobic residues. In at least half of these cases, however, either the relative position of the acidic or hydrophobic residues are different; either change is likely to alter the interaction of the phosphorylation site with upstream kinases or downstream SH2-proteins, effectively changing the signal. The comparative association of IRS-1 and IRS-2 with various SH2 domains observed in our experiments is consistent with these predictions.
The mIRS-2 gene lies near the type 1 procollagen 4a locus (Col4al), proximal to the centromere on mouse chromosome 8 (37, 38). We have compared our interspecific map of chromosome 8 with a composite mouse linkage map that reports the map location of many uncloned mouse mutations (provided from the Mouse Genome Database, maintained by the Jackson Laboratory, Bar Harbor, ME). Irs2 maps in a region that lacks mouse mutations that might be expected for an alteration in this locus (data not shown). By contrast, Irs1 is located on mouse chromosome 1, and the human gene is on human chromosome 2q36-37 (39). A few point mutations have been found in the human IRS-1 gene, but their effect on function is unclear (40, 41). It will be important to determine whether mutations in the human IRS-2 gene contribute to insulin resistance of non-insulin-dependent diabetes mellitus or to immune system disorders. Toward this end, we have isolated a partial clone with a high degree of identity to murine IRS-2, which may be the human counterpart (M. F. White and D. Bernal, unpublished data).
The proximal region of mouse chromosome 8 shares regions of homology with human chromosomes 8p, 13q, and 19p. In particular, Col4al, the locus most closely linked with Irs2, has been placed on human 13q34, suggesting that Irs2 may reside there as well. However, it is interesting that Irs2 lies close to the murine insulin receptor. The Drosophila insulin receptor contains a COOH-terminal extension of 400 residues with similarity to the first half of the COOH-terminus of IRS-2 (42). It is possible that the evolutionarily early insulin receptor contained such a tail, and that during the evolution of higher organisms, this tail (which remains part of the Drosophila receptor) separated from the receptor and was reassembled into the IRS-2 gene. Based on the proximity and functional relation between the insulin receptor and IRS-2, hIrs2 may be adjacent to the human insulin receptor on chromosome 19.
Broad distribution of IRS proteins and their interaction with various receptor systems suggests that they are essential for maintaining cell survival and growth and normal metabolic regulation (9, 11). However, our finding that phosphorylated IRS-1, but not IRS-2, increases during differentiation of 3T3-L1 fibroblasts into adipocytes suggest that IRS-1 and IRS-2 may not mediate identical signals. Because IRS-1 predominates in adipocytes, it may best mediate metabolic effects of insulin in this cell context, including the stimulation of glucose transport and inhibition of lipolysis. The predominance of IRS-2 in hematopoietic cells suggests that it may be preferred by cytokine receptors and be best adapted to mediate mitogenesis. This hypothesis is consistent with our previous finding that IRS-2 mediates a very sensitive mitogenic response to IL-4 (21). Thus, although IRS-1 and IRS-2 may be serving redundant roles in some circumstances, they perform physiologic functions in several instances based upon tissue distribution. Not only are IRS-1 and IRS-2 differentially expressed, but our analysis using various SH2 proteins to probe phosphorylation site characteristics suggests that IRS-1 and IRS-2 are phosphorylated differently and may recruit alternate downstream SH2-proteins. Furthermore, the stimulating factor (IL-4 or insulin) alters the association of SH2 proteins with IRS-1 and IRS-2. Similar differences may also occur during interaction with other IL, IFN, or GH receptors.
Thus, signaling by IRS-1 and IRS-2 varies by 1) tissue type, 2) growth
factor, and 3) elements directed by the differing structures of the two
IRS proteins. The ability to switch between IRS-1 or IRS-2 in cell
context-dependent manner provides a unique mechanism for signal
diversity that would be absent from classic receptors that engage SH2
proteins directly at their autophosphorylation sites. While it is
interesting to note that the IRS-1(-/-) mouse displays
disordered growth and metabolism, no immune system dysfunction has been
detected, which is consistent with the general absence of IRS-1 from
hematopoietic cells. However, based on the expression of IRS-1 in
classical insulin target tissues, it is not surprising that IRS-2
cannot completely compensate for the absence of IRS-1 for carbohydrate
metabolism (22, 23). The relative importance of IRS-2 and IRS-1 for
growth and development and metabolic regulation awaits the generation
of IRS-2(-/-) mouse and direct comparison and mating with
the IRS-1(-/-) mouse. Moreover, the recent discovery that
IRS-1, but not IRS-2, may mediate the inhibitory effect of tumor
necrosis factor- (TNF) on the insulin receptor suggests that the
IRS proteins define and regulate the crossroads at which many diverse
signaling systems converge and diverge (43).
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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), and IRS-1 have
been described (6, 20). Bal-17 B lymphoma cells were maintained in
RPMI-1640 medium supplemented with 10% FBS, L-glutamine (2
mM), and 2-mercaptoethanol (50 µM). The
3T3-L1 cells were maintained in DMEM high-glucose supplemented with
10% bovine calf serum; 2 days post-confluence, differentiation was
induced by incubating the cells for 3 days in DMEM supplemented with
10% FBS, 5 µg/ml insulin, 1 µM dexamethasone. p19 cultures were performed as previously described (30). Briefly, undifferentiated p19 cells were grown on tissue culture quality plastic in DMEM-F12 containing 10% FCS. Differentiation of p19 cells was induced by plating in the presence of 1 µM retinoic acid for 4 days on bacterial quality dishes (to which cells do not attach). At the end of the fourth day the aggregates were decanted, plated on tissue culture quality plastic, and allowed to differentiate for 4 days.
32D cells or cells expressing the human receptors for insulin and IL-4 (32DIR/IL4R) were transfected with pCMVhisIRS-2 or pCMVhisIRS-1 by electroporation, as previously described (20, 45). Cells were selected by survival in 5 mM histidinol (Sigma), and expression of IRS-2 and IRS-1 was monitored by immunoblotting.
Epididymal adipocytes were isolated from Spraque-Dawley rats (280–300 g) using a modification of the original methods described by Rodbell (46). Fat was dissected and collected in modified Krebs Ringer bicarbonate (buffer A), supplemented with 10 mM HEPES (Sigma) 2.5% BSA (Sigma), and 200 nM adenosine (Sigma). The tissue was briefly minced, and then digested for 40 min in a 37 °C shaker bath set at 10 rpm/6 sec by adding 2 mg collagenase (Worthington Biochemical, Freehold, NJ) per gram of tissues (47). The digested tissue was passed through a 50-µm nylon screen and rinsed several times with buffer A to remove the collagenase, then rinsed several times with BSA-free buffer A. Equal volumes of adipocytes are treated without or with 80 nM insulin for 5 min with gentle rotation in a 37 °C water bath.
Southern Analysis of Mouse Genomic DNA
Genomic DNA (10 µg) from mouse liver (48) was digested
overnight with various restriction enzymes, resolved by
electrophoresis, and transferred to Hybond N membranes (Amersham,
Arlington Heights, IL) for Southern analysis. Two specific DNA probes,
which contain the sequences corresponding to approximately -2525 to
423 bp, and 2777 to 4468 bp of IRS-2, were obtained by digesting mG28
with KpnI. The KpnI fragments were isolated and
labeled with [32P]phosphate as previously described (48).
Hybridization was conducted overnight at 40 °C in 5 x saline
sodium citrate (SSC), 40% formamide, 5x Denhardt’s, 0.1% SDS, and
100 ng/ml of salmon sperm DNA. Final washing of the blots was at 65
°C in 0.5 x SSC containing 0.1% SDS.
Interspecific Mouse Backcross Mapping
Interspecific backcross progeny were generated by mating
(C57BL/6J x M. spretus)F1 females and
C57BL/6J males as described (49). A total of 205 N2 mice
were used to map the Irs2 locus. This mapping panel has been
typed for more than 2000 loci that are distributed among all of the
autosomes and the X chromosome (49). DNA isolation, restriction enzyme
digestion, agarose gel electrophoresis, Southern blot transfer, and
hybridization were performed as described (50). An approximately 300-bp
BamHI/EcoRI mouse cDNA fragment was labeled with
[32P]dCTP using a nick translation kit (Boeringer
Mannheim, Indianapolis, IN) and used as probe; the final wash
stringency was 0.5x saline sodium citrate phosphate, 0.1% SDS, 65
°C. The probe detected 13 kb and 20 kb BamHI fragments
from C57B1/6J and M. spretus DNA, respectively. The 20-kb
M. spretus-specific fragment was followed in backcross
mice.
A description of the probes and RFLPs for the linked insulin receptor
(Insr) and tissue plaminogen activator (Plat)
loci has previously been reported (37). The probe for type IV 1
procollagen (Col4al) has not previously been reported; it
was a 1.9-kb EcoRI fragment which detected 9.7 (C57BL/6J)
and 8.6 (M. spretus) kb XbaI fragments.
Recombination distances were calculated as described (51) using the
SPRETUS MADNESS program. Gene order was determined by minimizing the
number of recombination events required to explain the allele
distribution patterns.
Immunoprecipitation and Immunoblot Analysis
Quiesent cells were incubated for 1 min in the absence or
presence of 100 nM insulin or 10 nM IL-4, and
lysed in homogenization buffer. The lysates were incubated with
polyclonal antibodies, and the immune complexes were collected on
protein A and washed three times with homogenization buffer, denatured,
separated by 7.5% SDS-PAGE, and transferred to nitrocellulose
membranes (Schleicher & Schuell, Keene, NH) for immunoblotting (52).
The antibodies were prepared in rabbits (HRP Corp., Denver, PA) as
previously described (10). The IRS-2 was obtained using a GST-fusion
protein containing residues 619-746 of mouse IRS-2 (25) as antigen;
IRS-1 was obtained using recombinant rat IRS-1 purified from Sf9
cells infected with a recombinant baculovirus as antigen (25);
IRS-1CT (residues 1221–1234 of rat IRS-1) were made
with synthetic peptides coupled to Keyhole limpet hemocyanin (53).
PY antibodies were rabbit polyclonal (54) or mouse monoclonal 4G10
(UBI, Lake Placid, NY). p85 antibodies were purchased from UBI.
Differential Binding of SH2 Domains with the IRS-Proteins
GST-fusion proteins containing nSH2p85,
SH2fyn, SH2Grb2, nSH2PLC,
nSH2SHPTP2, SH2abl, and SH2Crk were
prepared as previously described (25). Cell lysates were prepared from
unstimulated, insulin-stimulated, or IL-4-stimulated cells
(32DIR,IL4R, 32DIR,IL4R/IRS-1 and
32DIR,IL4R/IRS-2 cells) in homogenization buffer. The
extracts were clarified by centrifugation at 100,000 x
g for 1 h at 4 °C. The supernatants were incubated
with the 1 µg GST fusion proteins containing SH2 domains as indicated
at 4 °C for 1 h and precipitated with glutathione Sepharose at 4
°C for 1 h, washed twice with 50 mM Tris-HCl (pH
7.4) containing 100 mM NaCl, 250 µg/ml BSA, 0.2
mM vanadate, and 0.4 mM
phenylmethylsulfonylfluoride, and boiled for 5 min in 100 µl of
Laemmli sample buffer containing 0.1 M dithiothreitol.
Samples were separated on 7.5% SDS-PAGE and analyzed by immunoblotting
(25, 52).
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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This work was supported by NIH Grants DK-38712 and DK-43808 (M.F.W.) and by the National Cancer Institute, Department of Health and Human Services under contract with ABL (N.A.J.). X.J.S. is a recipient of the Juvenile Diabetes Foundation, and M.G.M., Jr. was partially supported by the Medical Scientist Training Program at Harvard Medical School.
1 Present address: University of Vermont, College of Medicine, Given
C350, Burlington, Vermont 05405. 
Received for publication August 1, 1996. Revision received October 23, 1996. Accepted for publication November 6, 1996.
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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 G. SESTI, M. FEDERICI, M. L. HRIBAL, D. LAURO, P. SBRACCIA, and R. LAURO Defects of the insulin receptor substrate (IRS) system in human metabolic disorders FASEB J, October 1, 2001; 15(12): 2099 - 2111. [Abstract] [Full Text] [PDF]
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R. G. Richards, D. M. Klotz, M. R. Bush, D. K. Walmer, and R. P. DiAugustine E2-Induced Degradation of Uterine Insulin Receptor Substrate-2: Requirement for an IGF-I-Stimulated, Proteasome-Dependent Pathway Endocrinology, September 1, 2001; 142(9): 3842 - 3849. [Abstract] [Full Text] [PDF]
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A. Takano, I. Usui, T. Haruta, J. Kawahara, T. Uno, M. Iwata, and M. Kobayashi Mammalian Target of Rapamycin Pathway Regulates Insulin Signaling via Subcellular Redistribution of Insulin Receptor Substrate 1 and Integrates Nutritional Signals and Metabolic Signals of Insulin Mol. Cell. Biol., August 1, 2001; 21(15): 5050 - 5062. [Abstract] [Full Text] [PDF]
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 J. Zhang, J. Ou, Y. Bashmakov, J. D. Horton, M. S. Brown, and J. L. Goldstein Insulin inhibits transcription of IRS-2 gene in rat liver through an insulin response element (IRE) that resembles IREs of other insulin-repressed genes PNAS, March 16, 2001; (2001) 71054598. [Abstract] [Full Text]
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K. Tsuruzoe, R. Emkey, K. M. Kriauciunas, K. Ueki, and C. R. Kahn Insulin Receptor Substrate 3 (IRS-3) and IRS-4 Impair IRS-1- and IRS-2-Mediated Signaling Mol. Cell. Biol., January 1, 2001; 21(1): 26 - 38. [Abstract] [Full Text]
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L. Liang, J. Jiang, and S. J. Frank Insulin Receptor Substrate-1-Mediated Enhancement of Growth Hormone-Induced Mitogen-Activated Protein Kinase Activation Endocrinology, September 1, 2000; 141(9): 3328 - 3336. [Abstract] [Full Text] [PDF]
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T. Haruta, T. Uno, J. Kawahara, A. Takano, K. Egawa, P. M. Sharma, J. M. Olefsky, and M. Kobayashi A Rapamycin-Sensitive Pathway Down-Regulates Insulin Signaling via Phosphorylation and Proteasomal Degradation of Insulin Receptor Substrate-1 Mol. Endocrinol., June 1, 2000; 14(6): 783 - 794. [Abstract] [Full Text]
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F. Ahmad, L.-N. Cong, L. Stenson Holst, L.-M. Wang, T. Rahn Landstrom, J. H. Pierce, M. J. Quon, E. Degerman, and V. C. Manganiello Cyclic Nucleotide Phosphodiesterase 3B Is a Downstream Target of Protein Kinase B and May Be Involved in Regulation of Effects of Protein Kinase B on Thymidine Incorporation in FDCP2 Cells J. Immunol., May 1, 2000; 164(9): 4678 - 4688. [Abstract] [Full Text] [PDF]
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T. Uchida, M. G. Myers Jr., and M. F. White IRS-4 Mediates Protein Kinase B Signaling during Insulin Stimulation without Promoting Antiapoptosis Mol. Cell. Biol., January 1, 2000; 20(1): 126 - 138. [Abstract] [Full Text]
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V. R. Fantin, Q. Wang, G. E. Lienhard, and S. R. Keller Mice lacking insulin receptor substrate 4 exhibit mild defects in growth, reproduction, and glucose homeostasis Am J Physiol Endocrinol Metab, January 1, 2000; 278(1): E127 - E133. [Abstract] [Full Text] [PDF]
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 L. Poretsky, N. A. Cataldo, Z. Rosenwaks, and L. C. Giudice The Insulin-Related Ovarian Regulatory System in Health and Disease Endocr. Rev., August 1, 1999; 20(4): 535 - 582. [Abstract] [Full Text] [PDF]
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L. Liang, T. Zhou, J. Jiang, J. H. Pierce, T. A. Gustafson, and S. J. Frank Insulin Receptor Substrate-1 Enhances Growth Hormone-Induced Proliferation Endocrinology, May 1, 1999; 140(5): 1972 - 1983. [Abstract] [Full Text]
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O. V. Chaika, N. Chaika, D. J. Volle, H. Hayashi, Y. Ebina, L.-M. Wang, J. H. Pierce, and R. E. Lewis Mutation of Tyrosine 960 within the Insulin Receptor Juxtamembrane Domain Impairs Glucose Transport but Does Not Inhibit Ligand-mediated Phosphorylation of Insulin Receptor Substrate-2 in 3T3-L1 Adipocytes J. Biol. Chem., April 23, 1999; 274(17): 12075 - 12080. [Abstract] [Full Text] [PDF]
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F. Ahmad, G. Gao, L. M. Wang, T. R. Landstrom, E. Degerman, J. H. Pierce, and V. C. Manganiello IL-3 and IL-4 Activate Cyclic Nucleotide Phosphodiesterases 3 (PDE3) and 4 (PDE4) by Different Mechanisms in FDCP2 Myeloid Cells J. Immunol., April 15, 1999; 162(8): 4864 - 4875. [Abstract] [Full Text] [PDF]
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Y. Lu, J. Brush, and T. A. Stewart NSP1 Defines a Novel Family of Adaptor Proteins Linking Integrin and Tyrosine Kinase Receptors to the c-Jun N-terminal Kinase/Stress-activated Protein Kinase Signaling Pathway J. Biol. Chem., April 9, 1999; 274(15): 10047 - 10052. [Abstract] [Full Text] [PDF]
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L. Vaßen, W. Wegrzyn, and L. Klein-Hitpass Human Insulin Receptor Substrate-2 (IRS-2) Is a Primary Progesterone Response Gene Mol. Endocrinol., March 1, 1999; 13(3): 485 - 494. [Abstract] [Full Text]
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V. R. Fantin, B. E. Lavan, Q. Wang, N. A. Jenkins, D. J. Gilbert, N. G. Copeland, S. R. Keller, and G. E. Lienhard Cloning, Tissue Expression, and Chromosomal Location of the Mouse Insulin Receptor Substrate 4 Gene Endocrinology, March 1, 1999; 140(3): 1329 - 1337. [Abstract] [Full Text]
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L. Yenush, C. Zanella, T. Uchida, D. Bernal, and M. F. White The Pleckstrin Homology and Phosphotyrosine Binding Domains of Insulin Receptor Substrate 1 Mediate Inhibition of Apoptosis by Insulin Mol. Cell. Biol., November 1, 1998; 18(11): 6784 - 6794. [Abstract] [Full Text]
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H. Sakaue, W. Ogawa, M. Matsumoto, S. Kuroda, M. Takata, T. Sugimoto, B. M. Spiegelman, and M. Kasuga Posttranscriptional Control of Adipocyte Differentiation through Activation of Phosphoinositide 3-Kinase J. Biol. Chem., October 30, 1998; 273(44): 28945 - 28952. [Abstract] [Full Text] [PDF]
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E. L. K. Goh, T. J. Pircher, and P. E. Lobie Growth Hormone Promotion of Tubulin Polymerization Stabilizes the Microtubule Network and Protects Against Colchicine-Induced Apoptosis Endocrinology, October 1, 1998; 139(10): 4364 - 4372. [Abstract] [Full Text] [PDF]
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S. R. Bauer, M. J. Ruiz-Hidalgo, E. K. Rudikoff, J. Goldstein, and J. Laborda Modulated Expression of the Epidermal Growth Factor-Like Homeotic Protein dlk Influences Stromal-Cell-Pre-B-Cell Interactions, Stromal Cell Adipogenesis, and Pre-B-Cell Interleukin-7 Requirements Mol. Cell. Biol., September 1, 1998; 18(9): 5247 - 5255. [Abstract] [Full Text]
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G. Inoue, B. Cheatham, R. Emkey, and C. R. Kahn Dynamics of Insulin Signaling in 3T3-L1 Adipocytes. DIFFERENTIAL COMPARTMENTALIZATION AND TRAFFICKING OF INSULIN RECEPTOR SUBSTRATE (IRS)-1 AND IRS-2 J. Biol. Chem., May 8, 1998; 273(19): 11548 - 11555. [Abstract] [Full Text] [PDF]
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A. M. Valverde, M. Lorenzo, S. Pons, M. F. White, and M. Benito Insulin Receptor Substrate (IRS) Proteins IRS-1 and IRS-2 Differential Signaling in the Insulin/Insulin-Like Growth Factor-I Pathways in Fetal Brown Adipocytes Mol. Endocrinol., May 1, 1998; 12(5): 688 - 697. [Abstract] [Full Text]
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V. R. Fantin, J. D. Sparling, J. W. Slot, S. R. Keller, G. E. Lienhard, and B. E. Lavan Characterization of Insulin Receptor Substrate 4 in Human Embryonic Kidney 293 Cells J. Biol. Chem., April 24, 1998; 273(17): 10726 - 10732. [Abstract] [Full Text] [PDF]
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J. G. Jackson, M. F. White, and D. Yee Insulin Receptor Substrate-1 is the Predominant Signaling Molecule Activated by Insulin-like Growth Factor-I, Insulin, and Interleukin-4 in Estrogen Receptor-positive Human Breast Cancer Cells J. Biol. Chem., April 17, 1998; 273(16): 9994 - 10003. [Abstract] [Full Text] [PDF]
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A. M. Valverde, T. Teruel, P. Navarro, M. Benito, and M. Lorenzo Tumor Necrosis Factor-{alpha} Causes Insulin Receptor Substrate-2-Mediated Insulin Resistance and Inhibits Insulin-Induced Adipogenesis in Fetal Brown Adipocytes Endocrinology, March 1, 1998; 139(3): 1229 - 1238. [Abstract] [Full Text] [PDF]
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S.-O. Kim, J. Jiang, W. Yi, G.-S. Feng, and S. J. Frank Involvement of the Src Homology 2-containing Tyrosine Phosphatase SHP-2 in Growth Hormone Signaling J. Biol. Chem., January 23, 1998; 273(4): 2344 - 2354. [Abstract] [Full Text] [PDF]
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L. Zhou, H. Chen, C. H. Lin, L.-N. Cong, M. A. McGibbon, S. Sciacchitano, M. A. Lesniak, M. J. Quon, and S. I. Taylor Insulin Receptor Substrate-2 (IRS-2) Can Mediate the Action of Insulin to Stimulate Translocation of GLUT4 to the Cell Surface in Rat Adipose Cells J. Biol. Chem., November 21, 1997; 272(47): 29829 - 29833. [Abstract] [Full Text] [PDF]
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S. Sciacchitano and S. I. Taylor Cloning, Tissue Expression, and Chromosomal Localization of the Mouse IRS-3 Gene Endocrinology, November 1, 1997; 138(11): 4931 - 4940. [Abstract] [Full Text] [PDF]
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Y. Kaburagi, S. Satoh, H. Tamemoto, R. Yamamoto-Honda, K. Tobe, K. Veki, T. Yamauchi, E. Kono-Sugita, H. Sekihara, S. Aizawa, et al. Role of Insulin Receptor Substrate-1 and pp60 in the Regulation of Insulin-induced Glucose Transport and GLUT4 Translocation in Primary Adipocytes J. Biol. Chem., October 10, 1997; 272(41): 25839 - 25844. [Abstract] [Full Text] [PDF]
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T. C. Haddad and C. A. Conover Insulin and Interleukin-4 Induce Desensitization to the Mitogenic Effects of Insulin-like Growth Factor-I. PIVOTAL ROLE FOR INSULIN RECEPTOR SUBSTRATE-2 J. Biol. Chem., August 1, 1997; 272(31): 19525 - 19531. [Abstract] [Full Text] [PDF]
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E. L. K. Goh, T. Zhu, S. Yakar, D. LeRoith, and P. E. Lobie CrkII Participation in the Cellular Effects of Growth Hormone and Insulin-like Growth Factor-1. PHOSPHATIDYLINOSITOL-3 KINASE DEPENDENT AND INDEPENDENT EFFECTS J. Biol. Chem., June 2, 2000; 275(23): 17683 - 17692. [Abstract] [Full Text] [PDF]
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C. A. Aspinwall, W.-J. Qian, M. G. Roper, R. N. Kulkarni, C. R. Kahn, and R. T. Kennedy Roles of Insulin Receptor Substrate-1, Phosphatidylinositol 3-Kinase, and Release of Intracellular Ca2+ Stores in Insulin-stimulated Insulin Secretion in beta -Cells J. Biol. Chem., July 14, 2000; 275(29): 22331 - 22338. [Abstract] [Full Text] [PDF]
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J. Zhang, J. Ou, Y. Bashmakov, J. D. Horton, M. S. Brown, and J. L. Goldstein Insulin inhibits transcription of IRS-2 gene in rat liver through an insulin response element (IRE) that resembles IREs of other insulin-repressed genes PNAS, March 27, 2001; 98(7): 3756 - 3761. [Abstract] [Full Text] [PDF]
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P. M. Catalano, S. E. Nizielski, J. Shao, L. Preston, L. Qiao, and J. E. Friedman Downregulated IRS-1 and PPARgamma in obese women with gestational diabetes: relationship to FFA during pregnancy Am J Physiol Endocrinol Metab, March 1, 2002; 282(3): E522 - E533. [Abstract] [Full Text] [PDF]
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