ISOLATION OF A HEAT-LABILE PROTEIN FACTOR INVOLVED IN THE 3'-PROCESSING OF H4 PRE-MESSENGER-RNA

It is well documented that the snRNP particles play a role in the proc essing of histone pre-mRNAs. There is also evidence that in addition t o the snRNP particles, at least two more proteins are involved in the in vitro 3'-processing of histone pre-mRNAs. The aim of the present wo rk was the isolation and purification of one of these PF250 factors. P rocessing factor PF250 was isolated from HeLa nuclear extracts using i on exchange chromatography. Its ability to partially restore the proce ssing activity of heat inactivated nuclear extract was demonstrated in vitro using labelled H4 pre-mRNA. Our results indicate that nuclear e xtracts exposed to 50 degrees C for 15 min lacked any processing activ ity. Using a series of column chromatography purification steps we iso lated a thermolabile protein factor that partially restored the activi ty of heat inactivated HeLa cell nuclear extracts. It can be concluded that besides U7 snRNP particles at least one more thermolabile protei n factor is required for the maturation of H4 pre-mRNA in vitro. It ha s a molecular mass of 30 kDa and is inactivated by heating at 50 degre es C; These characteristics make it a likely candidate to be a regulat ory factor involved in pre-mRNA processing. Copyright (C) 1996 Elsevie r Science Ltd