Regulation of lens connexin 45.6 by apoptotic protease, caspase-3.
Gap junctions are important in maintaining lens homeostasis. Here we report that connexin 45.6 (Cx45.6) was partially truncated to a 46 kDa fragment during chicken lens development. This specific truncation initiated during embryonic days and the truncated fragment accumulated towards the later developmental stages. When membranes of the embryonic lens were subjected to caspase-3 treatment, the 46 kDa fragment of Cx45.6 was reproduced, suggesting apoptotic protease caspase-3 is a potential protease involved. The COOH-terminus of Cx45.6 in GST-fusion protein was also cleaved by caspase-3, confirming that Cx45.6 is a direct substrate of caspase-3. Induction of apoptosis in lens primary cultures regenerated the 46 kDa fragment and this cleavage was blocked by a caspase-3 inhibitor. Alteration of amino acid residue Asp364 or Glu367 to Ala prevented Cx45.6 from cleavage by caspase-3, suggesting that the cleavage site of Cx45.6 is likely to be between Glu367 and Gly361. Phosphorylation of Ser363, a known substrate for casein kinase II (CKII) in vivo, inhibited the cleavage of Cx45.6 by caspase-3. Thus, this study demonstrates that a lens connexin can be a direct target of caspase-3 and the cleavage by caspase-3 leads to the development-associated truncation of Cx45.6. Finally, caspase-3 mediated truncation can be modulated by the specific connexin phosphorylation.
Yin
X
X
Department of Biochemistry, University of Texas Health Science Center, San Antonio 78229-3900, USA.
Gu
S
S
Jiang
J X
JX
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D047390:Casein Kinase II; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D002642:Chick Embryo; D017630:Connexins / Q000235:genetics / Q000378:metabolism*; D007908:Lens, Crystalline / Q000737:chemistry / Q000196:embryology* / Q000378:metabolism*; D008954:Models, Biological; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012694:Serine / Q000378:metabolism
D017630:Connexins; D011993:Recombinant Fusion Proteins; D012694:Serine; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D053148:Caspase 3; D020169:Caspases