LAMIN DYNAMICS DURING SEA-URCHIN MALE PRONUCLEAR FORMATION IN-VITRO

The dynamics of lamin disassembly and reassembly during sea urchin mal e pronuclear development in vitro was investigated. Using five anti-la min antibodies, we monitored by immunofluorescence and immunoblotting the changes in lamins during sperm chromatin decondensation, nuclear e nvelope (NE) formation, and male pronuclear swelling in fertilized sea urchin egg cytoplasmic extracts. We report the existence of five prot eins in sperm nuclei and swollen male pronuclei (p49, p54, p65, p72, p 84) which react with the antibodies. The sperm antigens resist membran e permeabilization with lysolecithin and 0.1% Triton X (TX)-100, but a re removed from the lateral aspects of the nuclei by 1% TX-100. All fi ve are completely removed from nuclei within 10 min of incubation in e gg extracts. Initial chromatin decondensation and NE formation occur w ithout reassembly of the putative lamins, but all lamins or lamin epit ope-containing peptides assemble coordinately during pronuclear swelli ng promoted by adenosine 5'-triphospate. Of the five pronuclear antige ns, p49 and p54 appear to originate exclusively from the sperm. p65, p 72, and p84 are also present in the egg cytoplasm and may be contribut ed to pronuclei by either source. Assembly of putative lamins and nucl ear swelling, but not chromatin decondensation and nuclear envelope fo rmation, are prevented in lamin-depleted cytoplasmic extracts. Our res ults indicate that p49 and p54 are not necessary for nuclear swelling, cytoplasmic p65 is required for swelling, and p72 and p84 are by them selves not sufficient but may be involved with p65 in nuclear swelling and full pronuclear development in vitro. (C) 1995 Academic Press, In c.