<i>Arabidopsis</i> AtRRP44 has ribonuclease activity that is required to complement the growth defect of yeast <i>rrp44</i> Mutant

Abstract

The RNA exosome is a multiprotein complex responsible for 3′ to 5′ degradation and processing of various classes of RNAs in eukaryotes. Rrp44/Dis3 is the catalytic center of the RNA exosome in yeast and human. Previously, we identified Arabidopsis thaliana RRP44 (AtRRP44) as a single functional homolog of Rrp44/Dis3. Although AtRRP44 is potentially a catalytic center of the plant RNA exosome, the ribonuclease activity of AtRRP44 has not been tested. Here, we show that AtRRP44 has ribonuclease activity using in vitro translated recombinant proteins. Mutation of the aspartic acid residue D489 of AtRRP44 to asparagine (D489N) resulted in loss of ribonuclease activity, indicating that aspartic acid is at the active site. The wild-type AtRRP44 protein rescued the growth defect of Saccharomyces cerevisiae rrp44 mutants, but the D489N mutated AtRRP44 did not. This finding suggests that the ribonuclease activity of wild-type AtRRP44 is required for yeast cell viability. We also showed that AtRRP44 was highly expressed in organs experiencing active cell turnover, such as shoot apical meristem, root apical meristem, and lateral root primordium. Along with previous studies showing that loss of RRP44 in Arabidopsis is lethal, our results suggest that AtRRP44 has ribonuclease activity that is related to plant development.

An RNA exosome subunit mediates cell-to-cell trafficking of a homeobox mRNA via plasmodesmata

“…1K and fig. S2A) ( 25 – 27 ) in the vicinity of the RRP44 catalytic center (fig. S2B) ( 28 ), which supports the notion that they are important for function.…”

“…S7, A to D). We also assessed whether the trafficking function of AtRRP44A could be uncoupled from its RNA-processing activity by introducing a noncatalytic AtRRP44A mutant [AtRRP44A D489N ~RFP (D489N, Asp 489 →Asn)] ( 27 ) driven by its native promoter into atrrp44a mutant trichome rescue plants. This construct fully rescued KN1 trafficking and trichome formation (fig.…”