Fig. 4. smd1–1 cell extracts contain reduced levels of stable U1 and U5 snRNPs when Lhp1p is absent. (A–F) Extracts from wild-type (lane 1), SMD1 lhp1::LEU2 (lane 2) and smd1–1 cells containing either no LHP1 (lane 3), chromosomal LHP1 (lane 4) or plasmid LHP1 (lane 5) were fractionated in 4% polyacrylamide native gels and subjected to Northern analysis. Blots were probed to detect U1 snRNPs (A), U2 snRNPs (B), U4–containing snRNPs (C), U5-containing snRNPs (D), U6 snRNPs (E) and the Nme1 RNA-containing MRP (F). (G) Wild-type and smd1–1 lhp1::LEU2 cells were lysed in aqueous buffer as described in Materials and methods. The relative levels of U5 and Nme1 RNAs in the extract supernatants (lanes 3 and 4) are compared with the relative levels of these RNAs when the cells are lysed in hot phenol (lanes 1 and 2). Fractionation of the extract supernatants in native gels is shown in lanes 6, 7, 9 and 10. Lanes 5–7 are probed to detect U5 snRNPs and lanes 8–10 are probed to detect RNase MRP. To confirm that naked RNA remained on the gel, lanes 5 and 8 contain total RNA from wild-type cells. The band denoted by the asterisk may represent aggregated RNA, as the RNA was not heated prior to loading.