FIGURE 2.

Lsm10 and Lsm11 mutants fail to properly process histone pre-mRNA and are necessary for development. (A) Total RNA isolated from whole third instar larvae of the indicated genotypes was subjected to Northern analysis with a ³²P-labeled H3 probe. Note that the severity of the misprocessed H3 phenotype is similar in U7²⁰, Lsm10^G40E, and Lsm11^c02047 mutants, while the terminal developmental phenotype is different: viable for U7²⁰ and not viable for the Lsm10^G40E and Lsm11^c02047 null mutants. The Lsm10^f06616 hypomorph is viable. (B) Protein extracts prepared from embryos of the indicated genotypes were probed with anti-V5 antibodies by Western blotting. P[Lsm11⁺] is a transgene expressing a V5-Lsm11 with the endogenous Lsm11 promoter. “11” Refers to the homozygous Lsm11^c02047 genotype. Lane 3 contains protein from a nontransgenic control. Note that in a wild-type Lsm11 background there is very little accumulation of V5-Lsm11 protein. α–Tubulin is used as a loading control. (C) Protein extracts isolated from whole third instar larvae of the indicated genotypes were subjected to immunoprecipitation then Western blot analysis with anti-V5 antibody. Lane 2 contains protein from nontransgenic control. (D,E) RNA isolated from whole third instar larvae of the indicated genotypes was subjected to Northern analysis with ³²P-labeled H3 probe. “10” Refers to the Lsm10^G40E/Df mutant genotype. Note that there is very little misprocessed H3 in both Lsm11^c02047, P[Lsm11⁺] and Lsm10^G40E/Df, P[Lsm10⁺] genotypes. (F) Protein extracts prepared from whole third instar larvae of the indicated genotypes were probed with anti-Lsm10 antibodies. Note that the lane 1 genotype contains a single copy of P[Lsm10⁺], accounting for the reduction in Lsm10 accumulation relative to wild type (+). All other P[Lsm] transgenes are present in two copies.