Garcia-Gomez JJ, Lebaron S, Froment C, Monsarrat B, Henry Y, de la Cruz J

Spb4 is a putative ATP-dependent RNA helicase required for proper processing of 27SB pre-rRNAs and therefore for 60S ribosomal subunit biogenesis. To define the timing of association of this protein with pre-ribosomal particles, we have studied the composition of complexes that copurify with TAP-tagged Spb4. These complexes contain mainly the ...

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27SB pre-rRNAs and about 50 ribosome biogenesis proteins, primarily components of early pre-60S ribosomal particles. To a lesser extent, some protein factors of 90S pre-ribosomal particles and the 35S and 27SA pre-rRNAs also copurify with TAP-tagged Spb4. Moreover, we have obtained by site-directed mutagenesis an allele that results in the R360A substitution in the conserved motif VI of the Spb4 helicase domain. This allele causes a dominant negative phenotype when overexpressed in the wild-type strain. Cells expressing Spb4(R360A) display an accumulation of 35S and 27SB pre-rRNAs and a net 40S ribosomal subunit defect. TAP-tagged Spb4(R360A) displays a greater steady-state association with 90S pre-ribosomal particles than TAP-tagged wild-type Spb4. Together, our data indicate that Spb4 is a component of early nucle(ol)ar pre-60S ribosomal particles containing 27SB pre-rRNA. Apparently, Spb4 binds 90S pre-ribosomal particles and dissociates from pre-60S ribosomal particles after processing of 27SB pre-rRNA.

Date: Aug. 08, 2011

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