Two Putative RNA-Binding Proteins Function with Unequal Genetic Redundancy in the MOS4-Associated Complex      

Abstract

The MOS4-associated complex (MAC) is a highly conserved nuclear protein complex associated with the spliceosome. We recently purified the MAC from Arabidopsis (Arabidopsis thaliana) nuclei, identified its potential components by mass spectrometry, and showed that at least five core proteins in the MAC are required for defense responses in plants. Here, we report the characterization of a putative RNA-binding protein identified in the MAC named MAC5A and its close homolog MAC5B. We confirmed that MAC5A is a component of the MAC through coimmunoprecipitation with the previously described MAC protein CELL DIVISION CYCLE5 from Arabidopsis. In addition, like all other characterized MAC proteins, MAC5A fused to the Green Fluorescent Protein localizes to the nucleus. Double mutant analysis revealed that MAC5A and MAC5B are unequally redundant and that a double mac5a mac5b mutant results in lethality. Probably due to this partial redundancy, mac5a and mac5b single mutants do not exhibit enhanced susceptibility to virulent or avirulent pathogen infection. However, like other MAC mutations, mac5a-1 partially suppresses the autoimmune phenotypes of suppressor of npr1-1, constitutive1 (snc1), a gain-of-function mutant that expresses a deregulated Resistance protein. Our results suggest that MAC5A is a component of the MAC that contributes to snc1-mediated autoimmunity.

Alternative Splicing at the Intersection of Biological Timing, Development, and Stress Responses

“…Changes in AS of SNC1 in the mos4, cdc5, and mac3a mac3b mutants provide compelling evidence that MAC mediates AS of a subset of R genes (Xu et al, 2012a). Another predicted RBP copurifying with MOS4, MAC5A, is also involved in pathogen defense (Monaghan et al, 2010). The human MAC5A counterpart is RBM22 (for RNA Binding Motif Protein22) that interacts with U6 snRNA and pre-mRNAs and participates in splicing.…”

The evolutionarily conserved MOS4-associated complex

“…Like MOS4, AtCDC5 appears to be necessary for resistance mediated by a variety of CC-and TIR-NB-LRR R proteins, including SNC1, as well as for basal resistance [20]. AtCDC5 was found to associate in planta with MOS4 and PRL1 [20], MAC3 [21], and MAC5 [22].…”

“…MOS4 has been shown to associate with 24 proteins, with nineteen sharing homology to yeast and human proteins that are either components of or associated with the conserved spliceosome-associated complex [21]. The core of the MAC is composed of the Myb-transcription factor AtCDC5, the WD-40 protein PRL2, MOS4, and the U-box E3 ubiquitin ligase proteins MAC3A/MAC3B, while a variety of other proteins including the RNAbinding proteins MAC5A/MAC5B associate with this core (Table 1) [21,22]. The orthologous protein components of the NTC, CDC5L complex, and MAC indicate that this complex is highly conserved amongst eukaryotes, and in all three systems the complex seems to have a close association with the spliceosome.…”

Plant defense responses against viral and bacterial pathogen infections. Focus on RNA-binding proteins (RBPs)

“…They are highly homologous one to other and both have an RRM as well as a CCCH-type zinc finger domain [6]. In order to assess the contribution of these proteins in plant immunity, the T-DNA insertional snc1 mac5a and snc1 mac5b double mutants were used.…”

“…Both PTI and ETI induce downstream defense genes followed by RNA processing steps such as 3' polyadenylation, splicing, 5' capping, and mRNA export out of the nucleus. MAC5A/B bind to the MOS4-associated Complex (MAC), which binds to the spliceosome, what suggests the contribution of MAC5A/B to mRNA splicing during plant defense against pathogens [6]. In case of viral pathogens after entering the plant cell via cell wall, disrupted by insect damage or wounding, they uncoat and disassemble.…”