A discrete 10-kDa polypeptide (10K) is expressed from early stages in the embryonic chicken lens. Since this has potential as a marker for lens cell development, chicken 10K and its homologues from mouse and human lenses were identified by protein sequencing and cloning. Surprisingly, lens 10K proteins appear to be identical to a lymphokine, macrophage migration inhibitory factor (MIF), originally identified in activated human T cells. Using microdissection and PCR techniques, we find that expression of 10K/MIF is strongly correlated with cell differentiation in the developing chicken lens. Northern blot analysis shows that 10K/MIF is widely expressed in mouse tissues. These results suggest that proteins with MIF activity may have roles beyond the immune system, perhaps as intercellular messengers or part of the machinery of differentiation itself. Indeed, partial sequence of other small lens proteins identifies another MIF-related protein (MRP8) in calf lens. The relatively abundant expression of MIF in lens may have clinical significance, with the possibility of involvement in ocular inflammations that may follow damage to the lens.

We have developed a system using explanted embryonic chicken lens epithelia to express foreign recombinant genes containing crystallin DNA regulatory sequences introduced by calcium phosphate transfection. Optimal results were obtained with lens epithelia from 14-day embryos transfected 1 day after explantation and assayed 3 days later. When DNA sequences (-364 to

Cyclin-dependent kinases and their regulatory subunits, the cyclins, are known to regulate progression through the cell cycle. Yet these same proteins are often expressed in non-cycling, differentiated cells. This review surveys the available information about cyclins and cyclin-dependent kinases in differentiated cells and explores the possibility that these proteins may have important functions that are independent of cell cycle regulation.

Previous studies of the developing lens have shown that Notch signaling regulates differentiation of lens fiber cells by maintaining a proliferating precursor pool in the anterior epithelium. However, whether Notch signaling is further required after the onset of fiber cell differentiation is not clear. This work investigates the role of Notch2 and Jagged1 (Jag1) in secondary fiber cell differentiation using rat lens epithelial explants undergoing FGF-2 dependent differentiation in vitro. FGF induced Jag1 expression and Notch2 signaling (as judged by the appearance of activated Notch2 Intracellular Domain (N2ICD)) within 12–24 hours. These changes were correlated with induction of the Notch effector, Hes5, upregulation of N-cadherin (N-cad), and downregulation of E-cadherin (E-cad), a cadherin switch characteristic of fiber cell differentiation. Induction of Jag1 was efficiently blocked by U0126, a specific inhibitor of MAPK/ERK signaling, indicating a requirement for signaling through this pathway downstream of the FGF receptor. Other growth factors that activate MAPK/ERK signaling (EGF, PDGF, IGF) did not induce Jag1. Inhibition of Notch signaling using gamma secretase inhibitors DAPT and L-685, 458 or anti-Jag1 antibody markedly decreased FGF-dependent expression of Jag1 demonstrating Notch-dependent lateral induction. In addition, inhibition of Notch signaling reduced expression of N-cad, and the cyclin dependent kinase inhibitor, p57Kip2, indicating a direct role for Notch signaling in secondary fiber cell differentiation. These results demonstrate that Notch-mediated lateral induction of Jag1 is an essential component of FGF-dependent lens fiber cell differentiation.