Stress-inducible, murine protein mSTI1. Characterization of binding domains for heat shock proteins and in vitro phosphorylation by different kinases.
We have recently isolated the cDNA for the murine homologue of the stress-inducible phosphoprotein STI1 (also known as IEF SSP 3521 or p60). STI1 was previously shown to be 2-fold up-regulated in MRC-5 fibroblasts upon viral transformation and to exist in a macromolecular complex with heat shock proteins of the HSP 70 and 90 families. By peptide-sequencing we have identified the two heat shock proteins that bind to murine STI1 (mSTI1) as HSC 70 and HSP 84/86. We describe two separate binding regions within mSTI1 for the two heat shock proteins. In the presence of cell extracts, the N-terminal region of mSTI1 binds preferentially to HSC 70, whereas the C-terminal portion of the molecule promotes the binding of HSP 84/86. Heat treatment caused a strong induction of mSTI1 message without affecting the steady-state level of the protein significantly. In addition, heat treatment led to changes in the isoform-composition of mSTI1. pp70(s6k), pp90(rsk), and mitogen-activated protein kinase-activated protein kinase 2 were tested as possible STI1 kinases in vitro using recombinant mSTI1 as a substrate: only pp90(rsk) was able to phosphorylate recombinant mSTI1. In vitro kinase assays using casein kinase II suggest serine 189 to be a likely phosphorylation site in mSTI1.
Lässle
M
M
Department of Cell Biology and Surgery, Harvard Medical School and Children's Hospital, Boston, Massachusetts 02115, USA. lassle@mit.edu
Blatch
G L
GL
Kundra
V
V
Takatori
T
T
Zetter
B R
BR
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002457:Cell Extracts; D005656:Fungal Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D005982:Glutathione Transferase / Q000235:genetics; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction
D002457:Cell Extracts; D005656:Fungal Proteins; D018840:HSP70 Heat-Shock Proteins; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D011494:Protein Kinases