Abstract
Previously, we demonstrated that microinjection of phosphoinositide-specific phospholipase Cγy1 (PLCγ1) and lipase-defective mutants of PLCγ1 induced G0 growth arrested NIH 3T3 fibroblasts to enter S phase of the cell cycle. These experiments suggested that regions other than the catalytic domain of PLCγ1 may be responsible for inducing mitogenesis. To test other regions of PLCγ1 for DNA synthesis inducing activity, cDNA fragments encoding Src homology (SH) and pleckstrin homology (PH) domains were subcloned into the bacterial expression plasmid pGEX-2TK, and the GST fusion proteins were purified. The complete PLCγ1 SH domain peptide was found to induce DNA synthesis after microinjection into growth arrested fibroblasts. Peptides containing a single SH3 domain or two SH2 domains induced a partial response that was restored to full activity if they were co-injected. The PH domain peptide did not induce DNA synthesis. Thus, both SH3 and SH2 activity combine to give maximum DNA synthesis induction, demonstrating that non-catalytic structural domains of PLCγ1 have pronounced effects on mitogenic signaling pathways.
Original language | English |
---|---|
Pages (from-to) | 186-193 |
Number of pages | 8 |
Journal | Biochemical and Biophysical Research Communications |
Volume | 222 |
Issue number | 1 |
DOIs | |
State | Published - 6 May 1996 |
Bibliographical note
Funding Information:*Intramural Research Support Program, SAIC Frederick, NCI-Frederick Cancer Research and Development Center, Frederick, Maryland 21702; †Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, NIH, Bethesda, Maryland 20892; ‡Protein Engineering Group, KIBB, Korea Institute of Science and Technology, 52 Oun-Dong Yusong-Gu, Taejon 305-333, Korea; and §Laboratory of Biochemical Physiology, DBS, NCI-FCRDC, Frederick, Maryland 21702