Transcription efficiency of human polymerase III genes in vitro does not depend on the RNP-forming autoantigen La

Transcription of class III genes is conducted by multi-protein complexes co nsisting of polymerase III itself and several transcription factors. We est ablished a reconstituted in vitro transcription system from which the autoa ntigen La was removed by immunodepletion, This system showed no RNP formati on, but was still fully active in transcription. Supplementing such La-free transcription reactions with recombinant La restored the formation of La c omplexes with the newly synthesised RNA, but did not lead to enhanced trans cription efficiency. Furthermore, we developed a technique for the generati on and isolation of transcription complexes, assembled from purified transc ription factors and isolated by glycerol centrifugation. These complexes we re fully competent to re-initiate RNA synthesis but they were not associate d with La and their transcription rate could not be stimulated by addition of recombinant La. Therefore, we conclude that La does not act as a human p olymerase III transcription factor.