Studies of tyrosine phosphorylation and Src family tyrosine kinases in the lens epithelium.
OBJECTIVE
Tyrosine phosphorylation regulates many aspects of cell function; thus, cells that have different roles often display different patterns of tyrosine phosphorylation. Because there is interest in differential function of the anterior and equatorial regions of the lens epithelium, studies were conducted to compare tyrosine phosphorylation in the two zones.


METHODS
Anterior and equatorial regions of the porcine lens epithelium were collected. Using Western blot analysis, tissue homogenates were probed for tyrosine kinase proteins, phospho-Src, phosphotyrosine, and proliferating cell nuclear antigen (PCNA).


RESULTS
Phosphotyrosine immunoblots revealed a marked difference between the pattern of tyrosine phosphorylation in anterior and equatorial regions of the epithelium. Many more bands were detected in the equatorial region, and band density was greater. The abundance of total and active Src family kinases was higher at the equator than at the anterior epithelium. Src kinase activity, which was measured directly by quantifying phosphorylation of a synthetic target peptide using (32)P-gamma-ATP, was detectable only at the equator. In organ-cultured lenses, PP2, a specific inhibitor of the Src kinase family, reduced the density of the phosphotyrosine protein bands. The abundance of PCNA, a protein expressed in proliferating cells, also was reduced in PP2-treated lenses.


CONCLUSIONS
The results suggest that the higher Src family kinase activity at the equator contributes to the higher degree of protein phosphorylation observed in this region. The ability of PP2 to suppress PCNA expression suggests a possible link between the activity of Src family kinases and cell proliferation.
Tamiya
Shigeo
S
Department of Ophthalmology and Visual Sciences, University of Louisville, School of Medicine, KY 40202, USA.
Delamere
Nicholas A
NA
D000818:Animals; D015153:Blotting, Western; D049109:Cell Proliferation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004848:Epithelium / Q000201:enzymology; D007908:Lens, Crystalline / Q000201:enzymology*; D009924:Organ Culture Techniques; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors; D011743:Pyrimidines / Q000494:pharmacology; D011799:Quinazolines; D013552:Swine; D014443:Tyrosine / Q000378:metabolism*; D020032:Tyrphostins / Q000494:pharmacology; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
C412373:AG 1879; D004791:Enzyme Inhibitors; D018809:Proliferating Cell Nuclear Antigen; D011743:Pyrimidines; D011799:Quinazolines; D020032:Tyrphostins; C101044:RTKI cpd; D019000:Phosphotyrosine; D014443:Tyrosine; D019061:src-Family Kinases; D017027:Protein Tyrosine Phosphatases