RESUMO
Two types of UV-light-induced inhibitions of the synthesis of small nuclear RNA species U1, U2, U3, U4, and U5 were described previously: an immediate inhibition and a separate, delayed suppression that requires 1-2 hr of postirradiation cell incubation and UV doses that are about tenfold lower. In the present report, U1 RNA transcription in isolated nuclei from HeLa cells, assayed by RNAase T1 protection, reproduced the delayed inhibition. The sizes of the protected RNA fragments suggest that it is the initiation of U1 RNA transcription that is blocked during this inhibition. Transient expression of a marked human U1 RNA gene that contains 425 and 92 nucleotides of the 5' and 3' flanking sequences, respectively, showed delayed, but not immediate inhibition (while the endogenous U1 RNA genes exhibited immediate suppression). This indicates that continuity of the U1 gene flanking sequences beyond those segments and/or chromosomal integration of the U1 gene are not needed for the delayed inhibition, but may be required for the immediate inhibition. Irradiation of a U1 RNA gene, followed by its injection into Xenopus laevis oocyte nuclei, did not reproduce the immediate or delayed inhibitions. This suggests that direct UV radiation damage to DNA in the U1 RNA gene region is not the critical lesion in either the immediate or delayed UV-light-induced inhibitions of U1 RNA synthesis. In addition, the RNAase T1 protection pattern of transcripts synthesized in isolated nuclei from nonirradiated HeLa cells suggests that these cells may produce small amounts of U1 RNA molecules with variant nucleotide sequences in the mature region of the transcript.
