HISTOGRAPHIC RECORDING OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 (HIV-1)REGULATORY PROTEIN REV AND NUCLEAR FACTORS

HeLa cells and HeLa cells expressing the HIV-1 regulatory protein Rev were immunostained for Rev and pre-mRNA processing factors and examine d histographically by confocal laser scanning microscopy. Following sh ort pulse-labelling with bromouridine tri-phosphate nascent RNA gave a granular nucleoplasmic staining increasing somewhat towards the perip hery as did also the heterogeneous ribonucleoproteins (hnRNPs) A1 and particularly C1/C2, a distribution pattern which has not been describe d. The sm-antigen of the small ribonucleoprotein particle (snRNP) prot eins U1, U2, U4/U6 and U5 stained the nucleoplasm diffusely in additio n to speckles which co-localised with speckles of the non-snRNP splici ng factor SC-35. Brominated RNA and the hnRNPs A1 and C1/C2 were to va rying degrees excluded from the speckles. Rev concentrated in the nucl eolus and often as a perinucleolar ring/zone. Rev also stained the nuc leoplasm and cytoplasm without co-localising with the above-mentioned proteins or brominated RNA and was not enriched or excluded in SC-35 s peckles. The nucleolar proteins B23 and C23, like Rev, gave primarily a perinucleolar ring and stained the nucleoplasm but did not otherwise co-localise with Rev or with nuclear proteins. Histographic recording of immunofluorescence images proved to be a valuable tool in the stud y of localisation of HIV-1 Rev and cellular components and of possible co-localisations. A parallel comparison of the subcellular patterns o f pre-mRNA processing factors versus major nucleolar antigens is new a nd suggests that the factors are not strictly separated in the nucleop lasm.