Lens fiber cell differentiation, apoptosis and denucleation: Novel roles of NCOA6

Abstract

Lens fiber cell differentiation is characterized by cell elongation, accumulation of crystallins and other lens-specific proteins and coordinated degradation of subcellular organelles, including the nucleus. Destruction of organelles is essential to prevent light scattering. Degradation of nuclei (denucleation, karyolysis) during mammalian development is a special process limited to lens fiber cells, erythrocytes and keratinocytes. The molecular mechanisms that regulate lens fiber cell denucleation are poorly understood. Although denucleation and apoptosis are described as two distinct processes, recent data suggest that various components of the pro-apoptotic cellular machinery can be involved in normal lens fiber denucleation process.;Nuclear receptor coactivator 6 (NCOA6) is a multifunctional protein implicated in embryonic development, cell survival, and homeostasis. An 81-amino acid fragment, dnNCOA6, containing the N-terminal nuclear receptor box (LXXLL motif) of NCOA6, acts as a dominant-negative (dn) inhibitor of NCOA6. Here, we expressed dnNCOA6 in postmitotic transgenic mouse lens fiber cells. The transgenic lenses showed reduced growth; a wide spectrum of lens fiber cell differentiation defects, including reduced expression of gamma-crystallins; and cataract formation. Those lens fiber cells entered an alternate proapoptotic pathway, and the denucleation (karyolysis) process was stalled. Activation of caspase-3 at embryonic day (E) 13.5 was followed by double-strand breaks (DSBs) formation monitored via a biomarker, gamma-H2AX. Intense terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) signals were found at E16.5. Thus, a window of ∼72 h between these events suggested prolonged though incomplete apoptosis in the lens fiber cell compartment that preserved nuclei in its cells. Although the p53 tumor suppressor gene is not required for normal lens development including the denucleation, a large number of studies demonstrated specific roles of p53 to induce apoptosis in the lens fiber cell compartment. However, it is not clear whether pro-survival roles of Ncoa6 depend on the existence of p53. Genetic experiments showed that the apoptotic-like processes in the transgenic lens were both p53-dependent and p53-independent. Lens-specific deletion of Ncoa6 also resulted in disrupted lens fiber cell differentiation. Our data demonstrate a cell-autonomous role of Ncoa6 in lens fiber cell differentiation and suggest novel insights into the process of lens fiber cell denucleation and apoptosis.