Lamin dynamics during sea urchin male pronuclear formation in vitro

The dynamics of lamin disassembly and reassembly during sea urchin male pronuclear development in vitro was investigated. Using five anti-lamin antibodies, we monitored by immunofluorescence and immunoblotting the changes in lamins during sperm chromatin decondensation, nuclear envelope (NE) formation, and male pronuclear swelling in fertilized sea urchin egg cytoplasmic extracts. We report the existence of five proteins in sperm nuclei and swollen male pronuclei (p49, p54, p65, p72, p84) which react with the antibodies. The sperm antigens resist membrane permeabilization with lysolecithin and 0.1% Triton X (TX)-100, but are removed from the lateral aspects of the nuclei by 1% TX-100. All five are completely removed from nuclei within 10 min of incubation in egg extracts. Initial chromatin decondensation and NE formation occur without reassembly of the putative lamins, but all lamins or lamin epitope-containing peptides assemble coordinately during pronuclear swelling promoted by adenosine 5'-triphosphate. Of the five pronuclear antigens, p49 and p54 appear to originate exclusively from the sperm. p65, p72, and p84 are also present in the egg cytoplasm and may be contributed to pronuclei by either source. Assembly of putative lamins and nuclear swelling, but not chromatin decondensation and nuclear envelope formation, are prevented in lamin-depleted cytoplasmic extracts. Our results indicate that p49 and p54 are not necessary for nuclear swelling, cytoplasmic p65 is required for swelling, and p72 and p84 are by themselves not sufficient but may be involved with p65 in nuclear swelling and full pronuclear development in vitro.