DSP1 and DSP4 Act Synergistically in Small Nuclear RNA 3′ End Maturation and Pollen Growth

Abstract

Small nuclear RNAs (snRNAs) play essential roles in spliceosome assembly and splicing. Most snRNAs are transcribed by the DNAdependent RNA polymerase II (Pol II) and require 39-end endonucleolytic cleavage. We have previously shown that the Arabidopsis (Arabidopsis thaliana) Defective in snRNA Processing 1 (DSP1) complex, composed of at least five subunits, is responsible for snRNA 39 maturation and is essential for plant development. Yet it remains unclear how DSP1 complex subunits act together to process snRNAs. Here, we show that DSP4, a member of the metallo-b-lactamase family, physically interacts with DSP1 through its b-Casp domain. Null dsp4-1 mutants have pleiotropic developmental defects, including impaired pollen development and reduced pre-snRNA transcription and 39 maturation, resembling the phenotype of the dsp1-1 mutant. Interestingly, dsp1-1 dsp4-1 double mutants exhibit complete male sterility and reduced pre-snRNA transcription and 39-end maturation, unlike dsp1-1 or dsp4-1. In addition, Pol II occupancy at snRNA loci is lower in dsp1-1 dsp4-1 than in either single mutant. We also detected miscleaved pre-snRNAs in dsp1-1 dsp4-1, but not in dsp1-1 or dsp4-1. Taken together, these data reveal that DSP1 and DSP4 function is essential for pollen development, and that the two cooperatively promote pre-snRNA transcription and 39-end processing efficiency and accuracy. Small nuclear RNAs (snRNAs; Hadjiolov et al., 1966), a class of noncoding RNAs, are the basal components of the spliceosome and play essential roles in pre-mRNA splicing (Black et al., 1985; Ray et al., 1997; Guo et al., 2009). Their biogenesis involves transcription and subsequent processing steps. In Arabidopsis (Arabidopsis thaliana), the DNA-dependent RNA polymerase II (Pol II) synthesizes the primary snRNA transcripts (pre-snRNAs) U1, U2, U4, U5, and U12, but not U6, which is transcribed by Pol III (Carbon et al., 1987; Vankan and Filipowicz, 1988). Following transcription, pre-snRNAs are subjected to endonucleolytic cleavage at specific sites to remove the RNA fragment transcribed beyond the 39-end of mature snRNAs. In metazoans, snRNA 39-end maturation requires the Integrator Complex (INT; Baillat et al., 2005). INT contains at least 14 subunits (Baillat et al., 2005; Chen and Wagner, 2010), associates with the C-terminal domain of the largest subunit of Pol II, and depends on transcription for complex formation. It cotranscriptionally cleaves pre-snRNAs upstream of the required 39-box RNA motif (Uguen and Murphy, 2003, 2004; Baillat et al., 2005;Chen and Wagner, 2010). Integrator Subunit 11 (INT11) is a putative member of the metallob-lactamase (MBL)/b-CASP family of RNA endonucleases, is homologous to the cleavage and polyadenylation

Non-parametric GWAS: Another View on Genome-wide Association Study

“…Notably, LOC101209768 encoding tyrosine-protein phosphatase DSP1 is newly spotted. The lack of this gene would result in pleiotropic developmental defects including impaired pollen development, and thus it is related to flowering (Pu et al, 2019).…”

Alternative splicing of DSP1 enhances snRNA accumulation by promoting transcription termination and recycle of the processing complex

“…Alexander staining was performed as previously described (8). Pollen grain ultrastructures were observed using a TEM according to Pu et al (2019) (10). Briefly, anthers were fixed, dehydrated, embedded in Epon812, and cut to 80 nm sections with a diamond knife.…”

“…DSP1 acts as a scaffold to bridge CPSF73-I and its catalytically inactive homolog DSP4. DSP1 and DSP4 are both required for the efficient and accurate cleavage of presnRNAs (8,10). INT is composed of, at least, 14 subunits, including INT9, a paralog of CPSF73-I and its homolog INT11, a counterpart of DSP4.…”

Spliceosomal snRNAs, the Essential Players in pre‐mRNA Processing in Eukaryotic Nucleus: From Biogenesis to Functions and Spatiotemporal Characteristics

“…These findings indicate that DSP1 and DSP4 synergistically promote snRNA transcription and 3′ end maturation. [ 131 ] As a homolog of the metazoan Integrator subunit INT11, CPSF73‐I in Arabidopsis belongs to the endonuclease/β‐CASP family of RNA nucleases, which cleaves phosphodiester bonds in RNA. CPSF73‐I not only exists in the plant DSP complex and participates in the 3′ end maturation of pre‐snRNAs, but also interacts with CPSF100 and other proteins to form the CPSF complex, which cleaves the 3′ end of pre‐mRNAs and is involved in poly (A) tail formation.…”

“…In addition to its involvement in snRNA biogenesis, the Integrator complex also participates in the expression of other Pol II‐dependent RNA species, including non‐polyadenylated long non‐coding RNAs (lncRNAs), [ 132,133 ] PIWI‐interacting RNAs (piRNAs), [ 134 ] enhancer RNAs (eRNAs), [ 135,136 ] telomerase RNA (tertRNA) [ 131 ] and nascent pre‐mRNAs. [ 137 ] The expanded scope of noncoding RNAs and protein‐coding mRNAs targeted by Integrator is mainly mediated by Integrator's endonucleolytic activity, phosphatase activity through PP2A and Integrator‐PP2A association with Pol II.…”