Structural/functional properties of a mammalian multi-component structure containing all major spliceosomal small nuclear ribonucleoprotein particles.
An approx. 40 S multi-component structure, consisting of all major spliceosomal small nuclear ribonucleoprotein particles (snRNP) (U1, U2, U4/U6 and U5) in stable association with a large number of polypeptides, mainly in the range 50-210 kDa, has been reported to exist within rat liver nuclear extracts [Guialis, Moraitou, Patrinou-Georgoula and Dangli (1991) Nucleic Acids Res. 19, 287-296]. Using a new polyclonal antibody recognizing a 63 kDa protein component of the complex, this multi-snRNP assembly was detected within rat liver nuclear extracts as efficiently as with the antibody for the U2 snRNP-specific B' polypeptide. The 63 kDa protein was found to correspond to the 66 kDa subunit of the splicing factor SF3a, a known integral component of the HeLa 17 S U2 snRNP. Anti-2,2,7-trimethylguanosine affinity chromatography was an easy and efficient way of purifying the multi-snRNP complex from rat liver 40 S heterogeneous nuclear ribonucleoprotein particle (hnRNP)-containing sucrose gradient fractions. By subsequent glycerol-gradient sedimentation, all known snRNP forms active in RNA splicing were identified among its constituents. A complex structurally similar to the rat multi-snRNP was also identified in HeLa nuclear extracts. Preservation of hnRNP-snRNP interactions was observed within HeLa 40 S fractions. Moreover, these fractions were capable of restoring splicing activity when applied in reconstitution studies to supplement a micrococcal nuclease-treated splicing extract.
Moraitou
M
M
Institute of Biological Research and Biotechnology, The National Hellenic Research Foundation, 48 Vas. Constantinou Avenue, Athens 11635, Greece.
Patrinou-Georgoula
M
M
Guialis
A
A
D000818:Animals; D006367:HeLa Cells / Q000737:chemistry; D006801:Humans; D008099:Liver / Q000378:metabolism; D008322:Mammals; D008836:Micrococcal Nuclease / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry; D012322:RNA Precursors / Q000378:metabolism; D012326:RNA Splicing / Q000235:genetics; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000276:immunology; D051381:Rats; D017413:Ribonucleoprotein, U2 Small Nuclear; D017411:Ribonucleoproteins, Small Nuclear / Q000737:chemistry*; D017381:Spliceosomes / Q000737:chemistry*; D013329:Structure-Activity Relationship
D009687:Nuclear Proteins; D012322:RNA Precursors; D016601:RNA-Binding Proteins; D017413:Ribonucleoprotein, U2 Small Nuclear; D017411:Ribonucleoproteins, Small Nuclear; C088049:splicing factor 3a; D008836:Micrococcal Nuclease