PLC gamma 1 Src homology domain induces mitogenesis in quiescent NIH 3T3 fibroblasts.
Previously, we demonstrated that microinjection of phosphoinositide-specific phospholipase C gamma 1 (PLC gamma 1) and lipase-defective mutants of PLC gamma 1 induced G(0) growth arrested NIH 3T3 fibroblasts to enter S phase of the cell cycle. These experiments suggested that regions other than the catalytic domain of PLC gamma 1 may be responsible for inducing mitogenesis. To test other regions of PLC gamma 1 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 PLC gamma l 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 PLC gamma 1 have pronounced effects on mitogenic signaling pathways.
Smith
M R
MR
Intramural Research Support Program, NCI-Frederick Cancer Research and Development Center, Maryland 21702, USA.
Liu
Y L
YL
Kim
S R
SR
Bae
Y S
YS
Kim
C G
CG
Kwon
K S
KS
Rhee
S G
SG
Kung
H F
HF
D016475:3T3 Cells; D000818:Animals; D001798:Blood Proteins; D007527:Isoenzymes / Q000502:physiology*; D051379:Mice; D008938:Mitosis; D051966:Phospholipase C gamma; D010750:Phosphoproteins; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D010738:Type C Phospholipases / Q000502:physiology*; D018909:src Homology Domains
D001798:Blood Proteins; D007527:Isoenzymes; D010750:Phosphoproteins; C039077:platelet protein P47; D010738:Type C Phospholipases; D051966:Phospholipase C gamma