AN ULTRASTRUCTURAL CHARACTERIZATION OF IN VITRO-ASSEMBLED HNRNP C-PROTEIN RNA COMPLEXES

In mammalian cells approximately 700 nucleotide lengths of pre-mRNA ar e packaged during transcription by a unique group of abundant nuclear proteins to form a repeating array of regular ribonucleoprotein comple xes termed 30-40S heterogeneous nuclear ribonucleoprotein particles (h nRNP particles). We have used electron microscopy to examine complexes that form when in vitro-transcribed RNA is bound by one of the purifi ed native core-particle proteins which comprise the 40S monoparticle ( the C protein tetramer). Negatively stained images of the C protein te tramer bound to particle-length RNA (700 nt) demonstrate that three te tramers bind each RNA molecule to form a stable closed triangular comp lex. The triangular complexes have an isosceles shape with a base of 1 8.0 nm and sides of 23.0 nm. When RNA molecules of 230 nt are used as substrates single tetramers bind to form complexes that appear as smal l rounded structures with an average diameter of 9.7 nm. Twice this le ngth of RNA (456 nt) supports the assembly of mostly bilobed complexes that are 20.4 nm long and 11.8 nm wide. Images of the C protein-RNA c omplexes which assemble on 1400-nucleotide lengths of RNA (two particl e lengths of RNA) clearly show complexes composed of two triangles whi le three-triangle complexes are seen when 2100-nt lengths of RNA are u sed as the assembly substrate. These ultrastructural results demonstra te that groups of three C protein tetramers combine with the length of RNA packaged in monoparticles to form a discreet triad structure. Thi s interaction establishes a structural basis for the RNA length requir ement in monoparticle assembly and a mechanism for the packaging of na scent transcripts into a repeating array of regular structures. (C) 19 95 Academic Press, Inc.