Differential expression of the catalytic subunits for PP-1 and PP-2A and the regulatory subunits for PP-2A in mouse eye.

PURPOSE

Reversible protein phosphorylation is a fundamental regulatory mechanism in all biologic processes. Protein serine/threonine phosphatases-1 (PP-1) and 2A (PP-2A) account for 90% of serine/threonine phosphatase activity in eukaryote cells and play distinct roles in regulating multiple cellular processes and activities. Our previous studies have established the expression patterns of the catalytic subunits for PP-1 (PP-1cs) and PP-2A (PP-2Acs) in bovine and rat lenses. In the present study, we have determined the expression patterns of PP-1cs (PP-1alpha and PP-1beta) and PP-2Acs (PP-2Aalpha and PP-2Abeta) in the retina and cornea along with the ocular lens of the mouse eye. Moreover, since the function of PP-2A is largely relied on its regulatory subunits, we have also analyzed the expression patterns of the genes encoding the scaffold A subunits of PP-2A, PP2A-Aalpha and PP2A-Abeta, and the regulatory B family subunits of PP-2A, PP2A-Balpha, PP2A-Bbeta, and PP2A-Bgamma. In addition, we have also demonstrated the differential protections of PP-1 and PP-2A in mouse lens epithelial cell line, alphaTN4-1, against oxidative stress-induced apoptosis.

METHODS

Total RNAs and proteins were extracted from the retina, lens epithelium, lens fiber cells, and cornea of the mouse eye. Reverse transcription polymerase chain reaction (RT-PCR) and real time PCR were used to detect the mRNA expression. Western blot and immunohistochemistry analysis were applied to examine the protein expression and distribution. Stable clones of alphaTN4-1 cells expressing either PP-1alpha or PP-2Aalpha were used to analyze the differential protections against oxidative stress-induced apoptosis.

RESULTS

PP-1 is more abundant than PP-2A in the mouse eye. The catalytic subunits for PP-1 and PP-2A display similar expression patterns in the retina and cornea but much reduced in the lens. The mRNAs for all five isoforms of PP2A-A and PP2A-B subunits are highly expressed in the retina, but only three out of the five mRNAs are expressed in the cornea. In the ocular lens, only PP2A-Abeta and PP2A-Bgamma mRNAs are clearly detectable. The A and B subunit proteins of PP-2A are highly expressed in the retina and cornea but are much reduced in the ocular lens. PP2A-Aalpha/beta are differentially distributed in the mouse retina.When transfected into mouse lens epithelial cells, alphaTN4-1, PP-1alpha and PP-2Aalpha display differential protection against oxidative stress-induced apoptosis.

CONCLUSIONS

Our results lead to the following conclusions regarding PP-1 and PP-2A in mouse eye: 1) PP1 is a more abundant phosphatase than PP-2A; 2) both PP-1 and PP-2A may play important roles, and the functions of PP-2A appear to be highly regulated by various regulatory subunits; and 3) the genes encoding PP-1alpha/beta, PP-2Aalpha/beta, PP-2A-Aalpha/beta, and PP-2A-B alpha/beta/gamma are all differentially expressed.