Pm. Algenstaedt et al., INSULIN-RECEPTOR SUBSTRATE PROTEINS CREATE A LINK BETWEEN THE TYROSINE PHOSPHORYLATION CASCADE AND THE CA2-ATPASES IN MUSCLE AND HEART(), The Journal of biological chemistry, 272(38), 1997, pp. 23696-23702
Following phosphorylation by tile insulin receptor kinase, the insulin
receptor substrates (IRS)-1 and IRS-2 bind to and activate several Sr
c homology 2 (SH2) domain proteins. To identify novel proteins that in
teract with IRS proteins in muscle, a human skeletal muscle cDNA expre
ssion library was created in the lambda EXLox system and probed with b
aculovirus-produced and tyrosine-phosphorylated human IRS-I, One clone
of the 10 clones which was positive through three rounds of screening
represented the C terminus of the human homologue of the adult fast t
witch skeletal muscle Ca2+ ATPase (SERCA1) including the cytoplasmic t
ail and part of transmembrane region 10, Western blot analysis of extr
acts of rat muscle demonstrated co-immunoprecipitation of both IRS-1 a
nd IRS-2 with the skeletal muscle Ca2+-ATPase (SERCA1) and the cardiac
muscle isoform (SERCA2), In both cases, injection of insulin stimulat
ed a 2- to 6-fold increase in association of which was maximal within
5 min, In primary cultures of aortic smooth muscle cells and C2C12 cel
ls, the insulin-stimulated interaction between IRS proteins and SERCA1
and -2 was dose-dependent with a maximum induction at 100 nM insulin,
This interaction was confirmed in a ''pull down'' experiment using a
glutathione S-transferase fusion protein containing the C terminus of
the human SERCA. isoform and phosphorylated IRS-I in vitro and could b
e blocked by a FLVRES-like domain peptide present in the human SERCA s
equence, Affinity chromatography of phosphopeptide libraries using the
glutathione S-transferase fusion protein of the C terminus of SERCA1
indicated a consensus sequence for binding of XpYGSS; this is identica
l to potential tyrosine phosphorylation sites at position 431 of human
IRS-1 and at position 500 of human IRS-2, In streptozotocin diabetic
rats the interaction between IRS proteins and SERCA1 in skeletal muscl
e and SERCA2 in cardiac muscle was significantly reduced, Taken togeth
er, these results indicate that the IRS proteins bind to the Ca2+-ATPa
se of the sarcoplasmic reticulin in an insulin-regulated fashion, thus
creating a potential link between the tyrosine phosphorylation cascad
e and effects of insulin on calcium.