Crystallins and their synthesis in human lens epithelial cells in tissue culture (original) (raw)
Related papers
Biochemical and Biophysical Research Communications, 1974
incorporation of ~3H3 leucine into ollgomeric a-crystallin via individual subunits has beenmeasured in epithelial cells and cortex fiber cells from adult bovine lenses in vitro. Our data show that the ratio of [3H~ leucine incorporation via subunlts aB2 and aA 2 Ts shifted from a value of about 1:2 in epithelial cells to a value of about 1:3 in fiber cells. Thus, in this system, cellular differentiation is accompanied by a change in the stolchiometry of assembly of individual subunits to form the oligomerlc a-crystallin molecule. These results indicate possible changes in the rates of synthesis of individual a-crystallln subunits. a-Crystallin is an ollgomerlc structural protein that is found in both epithelial cells and fiber cells of the bovine lens (1) and has a molecular weight of approximately 1 X 106 (2, 3). Treatment with urea and mercaptoethanol dissociates the oligomer into its polypeptide subunits, all of which have single molecular weights of about 25,000 (4). These subunits can be resolved into two acidic proteins (aA 1 and aA2) , which have isoelectric points of 5.6 and 5.9, respectively, and two basic proteins (aB 1 and aB2) , which have isoelectric points *Research sponsored by the U. S. Atomic Energy Commission under contract with the Union Carbide Corporation.
Reliability of delta-crystallin as a marker for studies of chick lens induction
Differentiation, 1998
Induction of a lens by the optic vesicle of the brain was the first demonstration of how tissue interactions could influence cell fate during development. However, recent work with amphibians has shown that the optic vesicle is not the primary inducer of lens formation. Rather, an earlier interaction between anterior neural plate and presumptive lens ectoderm appears to direct lens formation. One problem with many early experiments was the absence of an unambiguous assay for lens formation. Before being able to test whether the revised model of lens induction applies to chicken embryos, we examined the suitability of using delta-crystallin as a marker of lens formation. Although delta-crystallin is the major protein synthesized in the chick lens, one or both of the two delta-crystallin genes found in chickens is transcribed in many non-lens tissues as well. In studies of lens formation where appearance of the delta-crystallin protein is used as a positive assay, synthesis of deltacrystallin outside of the lens could make experiments difficult to interpret. Therefore, polyacrylamide gel electrophoresis, immunoblotting, and immunofluorescence were used to determine whether the delta-crystallin messenger RNA detected in non-lens tissues is translated into protein, as it is in the lens. On Coomassie-blue-stained gels of several tissues from stage-22 embryos, a prominent protein was observed that co-migrated with deltacrystallin. However, on immunoblots, none of the non-lens tissues tested contained detectable levels of deltacrystallin at this stage. By imunofluorescence, deltacrystallin was observed in Rathke's pouch and in a large area of oral ectoderm near Rathke's pouch, yet none of the cells in these non-lens tissues showed the typical elongated morphology of lens fiber cells. When presumptive lens ectoderm or other regions of ectoderm from stage-10 embryos were cultured and tested for lens differentiation, both cell elongation and delta-crystallin synthesis were observed, or neither were observed. The results suggest that delta-crystallin synthesis and cell elongation together serve as useful criteria for assessing a positive lens response.& b d y :