RNA Binding Proteins RZ-1B and RZ-1C Play Critical Roles in Regulating Pre-mRNA Splicing and Gene Expression during Development in Arabidopsis

Plant-specific RNA binding proteins RZ-1B and RZ-1C interact with a number of highly conserved serine/arginine-rich proteins to regulate RNA splicing and gene expression in Arabidopsis. Nuclear-localized RNA binding proteins are involved in various aspects of RNA metabolism, which in turn modulates gene expression. However, the functions of nuclear-localized RNA binding proteins in plants are poorly understood. Here, we report the functions of two proteins containing RNA recognition motifs, RZ-1B and RZ-1C, in Arabidopsis thaliana. RZ-1B and RZ-1C were localized to nuclear speckles and interacted with a spectrum of serine/arginine-rich (SR) proteins through their C termini. RZ-1C preferentially bound to purine-rich RNA sequences in vitro through its N-terminal RNA recognition motif. Disrupting the RNA binding activity of RZ-1C with SR proteins through overexpression of the C terminus of RZ-1C conferred defective phenotypes similar to those observed in rz-1b rz-1c double mutants, including delayed seed germination, reduced stature, and serrated leaves. Loss of function of RZ-1B and RZ-1C was accompanied by defective splicing of many genes and global perturbation of gene expression. In addition, we found that RZ-1C directly targeted FLOWERING LOCUS C (FLC), promoting efficient splicing of FLC introns and likely also repressing FLC transcription. Our findings highlight the critical role of RZ-1B/1C in regulating RNA splicing, gene expression, and many key aspects of plant development via interaction with proteins including SR proteins.

[1]  C. Dean,et al.  Quantitative regulation of FLC via coordinated transcriptional initiation and elongation , 2015, Proceedings of the National Academy of Sciences.

[2]  Alex P. Reynolds,et al.  Native Elongating Transcript Sequencing Reveals Human Transcriptional Activity at Nucleotide Resolution , 2015, Cell.

[3]  M. Ares,et al.  Context-dependent control of alternative splicing by RNA-binding proteins , 2014, Nature Reviews Genetics.

[4]  C. Dean,et al.  Antisense-mediated FLC transcriptional repression requires the P-TEFb transcription elongation factor , 2014, Proceedings of the National Academy of Sciences.

[5]  Caroline Dean,et al.  Functional Consequences of Splicing of the Antisense Transcript COOLAIR on FLC Transcription , 2014, Molecular cell.

[6]  Jigang Li,et al.  TRANSLUCENT GREEN, an ERF family transcription factor, controls water balance in Arabidopsis by activating the expression of aquaporin genes. , 2014, Molecular plant.

[7]  D. Bentley Coupling mRNA processing with transcription in time and space , 2014, Nature Reviews Genetics.

[8]  P. Cui,et al.  A KH-Domain RNA-Binding Protein Interacts with FIERY2/CTD Phosphatase-Like 1 and Splicing Factors and Is Important for Pre-mRNA Splicing in Arabidopsis , 2013, PLoS genetics.

[9]  H. Gu,et al.  Membrane-Bound RLCKs LIP1 and LIP2 Are Essential Male Factors Controlling Male-Female Attraction in Arabidopsis , 2013, Current Biology.

[10]  Charles Y. Lin,et al.  SR Proteins Collaborate with 7SK and Promoter-Associated Nascent RNA to Release Paused Polymerase , 2013, Cell.

[11]  Gene W. Yeo,et al.  Genome-wide analysis reveals SR protein cooperation and competition in regulated splicing. , 2013, Molecular cell.

[12]  C. Zipfel,et al.  Pseudomonas HopU1 modulates plant immune receptor levels by blocking the interaction of their mRNAs with GRP7 , 2013, The EMBO journal.

[13]  Dorothee Staiger,et al.  A Circadian Clock-Regulated Toggle Switch Explains AtGRP7 and AtGRP8 Oscillations in Arabidopsis thaliana , 2013, PLoS Comput. Biol..

[14]  C. Zipfel,et al.  Pseudomonas HopU 1 modulates plant immune receptor levels by blocking the interaction of their mRNAs with GRP 7 , 2013 .

[15]  A. Ben-Hur,et al.  Identification of an intronic splicing regulatory element involved in auto-regulation of alternative splicing of SCL33 pre-mRNA. , 2012, The Plant journal : for cell and molecular biology.

[16]  Z. Weng,et al.  The Cellular EJC Interactome Reveals Higher-Order mRNP Structure and an EJC-SR Protein Nexus , 2012, Cell.

[17]  D. Black,et al.  Transcript Dynamics of Proinflammatory Genes Revealed by Sequence Analysis of Subcellular RNA Fractions , 2012, Cell.

[18]  H. Urlaub,et al.  Crystal structure of Cwc2 reveals a novel architecture of a multipartite RNA‐binding protein , 2012, The EMBO journal.

[19]  S. Jacobsen,et al.  The splicing factor SR45 affects the RNA-directed DNA methylation pathway in Arabidopsis , 2012, Epigenetics.

[20]  H. Urlaub,et al.  Crystal structure of Cwc 2 reveals a novel architecture of a multipartite RNA-binding protein , 2012 .

[21]  M. Rosbash,et al.  Nascent-seq indicates widespread cotranscriptional pre-mRNA splicing in Drosophila. , 2011, Genes & development.

[22]  S. Stamm,et al.  An SRp75/hnRNPG complex interacting with hnRNPE2 regulates the 5' splice site of tau exon 10, whose misregulation causes frontotemporal dementia. , 2011, Gene.

[23]  Caihong Yu,et al.  CFL1, a WW Domain Protein, Regulates Cuticle Development by Modulating the Function of HDG1, a Class IV Homeodomain Transcription Factor, in Rice and Arabidopsis[W] , 2011, Plant Cell.

[24]  Minami Matsui,et al.  A high-throughput screening system for Arabidopsis transcription factors and its application to Med25-dependent transcriptional regulation. , 2011, Molecular plant.

[25]  Ji Hoon Han,et al.  Structural determinants crucial to the RNA chaperone activity of glycine-rich RNA-binding proteins 4 and 7 in Arabidopsis thaliana during the cold adaptation process , 2011, Journal of experimental botany.

[26]  Jingdong Tian,et al.  Circular polymerase extension cloning for high-throughput cloning of complex and combinatorial DNA libraries , 2011, Nature Protocols.

[27]  Peng Cui,et al.  Arginine methylation mediated by the Arabidopsis homolog of PRMT5 is essential for proper pre-mRNA splicing , 2010, Proceedings of the National Academy of Sciences.

[28]  J. Y. Kim,et al.  Comparative analysis of Arabidopsis zinc finger-containing glycine-rich RNA-binding proteins during cold adaptation. , 2010, Plant physiology and biochemistry : PPB.

[29]  A. Barta,et al.  Implementing a Rational and Consistent Nomenclature for Serine/Arginine-Rich Protein Splicing Factors (SR Proteins) in Plants , 2010, Plant Cell.

[30]  J. Y. Kim,et al.  Glycine-rich RNA-binding proteins are functionally conserved in Arabidopsis thaliana and Oryza sativa during cold adaptation process , 2010, Journal of experimental botany.

[31]  Xuegong Zhang,et al.  DEGseq: an R package for identifying differentially expressed genes from RNA-seq data , 2010, Bioinform..

[32]  Yong Guo,et al.  Dof5.6/HCA2, a Dof Transcription Factor Gene, Regulates Interfascicular Cambium Formation and Vascular Tissue Development in Arabidopsis[W][OA] , 2009, The Plant Cell Online.

[33]  Z. Lorković,et al.  Role of plant RNA-binding proteins in development, stress response and genome organization. , 2009, Trends in plant science.

[34]  Lior Pachter,et al.  Sequence Analysis , 2020, Definitions.

[35]  J. Cáceres,et al.  The SR protein family of splicing factors: master regulators of gene expression. , 2009, The Biochemical journal.

[36]  C. Pikaard,et al.  Noncoding Transcription by RNA Polymerase Pol IVb/Pol V Mediates Transcriptional Silencing of Overlapping and Adjacent Genes , 2008, Cell.

[37]  D. Staiger,et al.  The small glycine-rich RNA binding protein AtGRP7 promotes floral transition in Arabidopsis thaliana. , 2008, The Plant journal : for cell and molecular biology.

[38]  Hwa Jung Lee,et al.  Glycine-rich RNA-binding protein 7 affects abiotic stress responses by regulating stomata opening and closing in Arabidopsis thaliana. , 2008, The Plant journal : for cell and molecular biology.

[39]  B. Williams,et al.  Mapping and quantifying mammalian transcriptomes by RNA-Seq , 2008, Nature Methods.

[40]  Yuxian Zhu,et al.  Mutation of Arabidopsis BARD1 Causes Meristem Defects by Failing to Confine WUSCHEL Expression to the Organizing Center[W][OA] , 2008, The Plant Cell Online.

[41]  S. Tabata,et al.  The Arabidopsis OBERON1 and OBERON2 genes encode plant homeodomain finger proteins and are required for apical meristem maintenance , 2008, Development.

[42]  K. Uchida,et al.  Auto-regulation of the circadian slave oscillator component AtGRP7 and regulation of its targets is impaired by a single RNA recognition motif point mutation. , 2007, The Plant journal : for cell and molecular biology.

[43]  Sandra Pelletier,et al.  A Receptor-like Kinase Mediates the Response of Arabidopsis Cells to the Inhibition of Cellulose Synthesis , 2007, Current Biology.

[44]  A. Reddy,et al.  Regulation of Plant Developmental Processes by a Novel Splicing Factor , 2007, PloS one.

[45]  Sung-ju Ahn,et al.  Functional characterization of a glycine-rich RNA-binding protein 2 in Arabidopsis thaliana under abiotic stress conditions. , 2007, The Plant journal : for cell and molecular biology.

[46]  K. Hibara,et al.  Arabidopsis CUP-SHAPED COTYLEDON3 Regulates Postembryonic Shoot Meristem and Organ Boundary Formation[W] , 2006, The Plant Cell Online.

[47]  J. Callis Faculty Opinions recommendation of An indole-3-acetic acid carboxyl methyltransferase regulates Arabidopsis leaf development. , 2005 .

[48]  Hunseung Kang,et al.  Cold-inducible zinc finger-containing glycine-rich RNA-binding protein contributes to the enhancement of freezing tolerance in Arabidopsis thaliana. , 2005, The Plant journal : for cell and molecular biology.

[49]  Volker Brendel,et al.  The ASRG database: identification and survey of Arabidopsis thaliana genes involved in pre-mRNA splicing , 2004, Genome Biology.

[50]  Ruthie Angelovici,et al.  Detection of protein-protein interactions in plants using bimolecular fluorescence complementation. , 2004, The Plant journal : for cell and molecular biology.

[51]  T. S. Wang,et al.  The B-Subunit of DNA Polymerase α-Primase Associates with the Origin Recognition Complex for Initiation of DNA Replication , 2004, Molecular and Cellular Biology.

[52]  S. Rhee,et al.  MAPMAN: a user-driven tool to display genomics data sets onto diagrams of metabolic pathways and other biological processes. , 2004, The Plant journal : for cell and molecular biology.

[53]  M. Sugiura,et al.  Isolation of a novel RNA-binding protein and its association with a large ribonucleoprotein particle present in the nucleoplasm of tobacco cells , 1996, Plant Molecular Biology.

[54]  A. Barta,et al.  Ectopic expression of atRSZ33 reveals its function in splicing and causes pleiotropic changes in development. , 2003, Molecular biology of the cell.

[55]  S. D. de Vries,et al.  The CUP-SHAPED COTYLEDON3 Gene Is Required for Boundary and Shoot Meristem Formation in Arabidopsis Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.012203. , 2003, The Plant Cell Online.

[56]  D. Inzé,et al.  DRL1, a Homolog of the Yeast TOT4/KTI12 Protein, Has a Function in Meristem Activity and Organ Growth in Plants Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.007062. , 2003, The Plant Cell Online.

[57]  A. Barta,et al.  Network of Interactions of a Novel Plant-specific Arg/Ser-rich Protein, atRSZ33, with atSC35-like Splicing Factors* , 2002, The Journal of Biological Chemistry.

[58]  Andrea Barta,et al.  Genome analysis: RNA recognition motif (RRM) and K homology (KH) domain RNA-binding proteins from the flowering plant Arabidopsis thaliana. , 2002, Nucleic acids research.

[59]  A. Reddy,et al.  An SC35-like Protein and a Novel Serine/Arginine-rich Protein Interact with Arabidopsis U1-70K Protein* , 1999, The Journal of Biological Chemistry.

[60]  H. Kimura,et al.  Quantitation of RNA Polymerase II and Its Transcription Factors in an HeLa Cell: Little Soluble Holoenzyme but Significant Amounts of Polymerases Attached to the Nuclear Substructure , 1999, Molecular and Cellular Biology.

[61]  A. Krainer,et al.  atSRp30, one of two SF2/ASF-like proteins from Arabidopsis thaliana, regulates splicing of specific plant genes. , 1999, Genes & development.

[62]  J. Stévenin,et al.  The splicing factors 9G8 and SRp20 transactivate splicing through different and specific enhancers. , 1999, RNA.

[63]  S. Hake,et al.  KNAT1 induces lobed leaves with ectopic meristems when overexpressed in Arabidopsis. , 1996, The Plant cell.

[64]  F. Nagy,et al.  Brassinosteroids Rescue the Deficiency of CYP90, a Cytochrome P450, Controlling Cell Elongation and De-etiolation in Arabidopsis , 1996, Cell.

[65]  U. Schibler,et al.  Physical isolation of nascent RNA chains transcribed by RNA polymerase II: evidence for cotranscriptional splicing , 1994, Molecular and cellular biology.