CAX-INTERACTING PROTEIN4 depletion causes early lethality and pre-mRNA missplicing in Arabidopsis.

Zinc knuckle (ZCCHC) motif-containing proteins are present in unicellular and multicellular eukaryotes, and most ZCCHC proteins with known functions participate in the metabolism of various classes of RNA, such as mRNAs, ribosomal RNAs, and microRNAs. The Arabidopsis (Arabidopsis thaliana) genome encodes 69 ZCCHC-containing proteins; however, the functions of most remain unclear. One of these proteins, CAX-INTERACTING PROTEIN 4 (CXIP4, encoded by AT2G28910), has been classified as a PTHR31437 family member. This family includes human Splicing regulatory glutamine/lysine-rich protein 1 (SREK1)-interacting protein 1 (SREK1IP1), which is thought to function in pre-mRNA splicing and RNA methylation. Metazoan SREK1IP1-like and plant CXIP4-like proteins only share a ZCCHC motif, and their functions remain almost entirely unknown. Here, we studied two loss-of-function alleles of Arabidopsis CXIP4: cxip4-1 is likely null and shows early lethality, and cxip4-2 is hypomorphic and viable, with pleiotropic morphological defects. The cxip4-2 mutant exhibited deregulation of defense genes and upregulation of transcription factor genes, some of which might explain its developmental defects. The cxip4-2 mutant also exhibited increased intron retention events, being more evident in cxip4-1. The specific functions of misspliced genes, such as those involved in "gene silencing by DNA methylation" and "mRNA polyadenylation factor" suggest that CXIP4 has additional functions. In cxip4-2 plants, polyadenylated RNAs accumulate in the nucleus; these could be misspliced mRNAs. The CXIP4 protein localizes to the nucleus in a pattern resembling nuclear speckles rich in splicing factors. Therefore, CXIP4 is required for plant development and survival and mRNA maturation.