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S. Tamiya, N.A. Delamere; Non–uniform expression and function of Src family tyrosine kinases in the porcine lens epithlium . Invest. Ophthalmol. Vis. Sci. 2004;45(13):1713.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: There is growing interest in differential function of the anterior and equatorial epithelium of the lens. Recently, we have shown Na,K–ATPase activity in the two regions is different (Tamiya et al., IOVS 44, 4395–99, 2003). Since the Na,K–ATPase function can be modulated by tyrosine phosphorylation (Bozulic et al., Am. J. Physiol. in press), studies were conducted to examine the expression and activity of Src kinase family. Methods: Lenses were dissected from fresh porcine eyes. The anterior and equatorial region of the epithelium were collected separately and homogenized in either RIPA buffer (Western blot) or Src kinase reaction buffer (Src kinase activity assay). For Western blots, homogenates were subjected to SDS–PAGE, transferred to nitrocellulose membranes, and probed for tyrosine kinase proteins, phospho–Src and phospho–tyrosine. Results: Phospho–tyrosine immunoblots revealed higher abundance of tyrosine phosphorylated proteins in the equatorial region of the epithelium compared to the anterior region. Judged by Western blot band density, the expression of Src kinase family members Src and Yes was higher at the equator compared to the anterior epithelium. Results from phospho–Src Western blots suggested that the active form of the Src kinase family is also higher at the equator. To confirm this finding, Src kinase activity was measured directly by quantifying phosphorylation of synthetic target peptides using 32P–γ–ATP. Src kinase family activity was higher at the equator compared to the anterior. Conclusions: Both the abundance and activity of the Src family of tyrosine kinases is higher at the equator compared to the anterior region. This finding is consistent with the possible role of Src in regulating lens cell proliferation at the equator. Further studies are required to determine whether Na,K–ATPase function in the equatorial epithelium is influenced by the activity of specific tyrosine kinases.
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