A repertoire of intronic lariat RNAs reveals tissue-specific regulation and target mimicry potential in plants

Lariat RNA is concomitantly produced by excised intron during RNA splicing,which is usually debranched by DBR1,an RNA debranching enzyme.However,increasing evidence showed that some lariat RNA could escape debranching.Little is known about how and why these lariat RNAs could be retained.By comparing the atlas of lariat RNAs between the non-dividing cell(mature pollen)and three actively dividing tissues(young shoot apex,young seeds,and young roots),we identified hundreds to thousands of lariat RNA naturally retained in each tissue,and the incidence of lariat RNA retention is much less in shoot apex while much more in pollen.Many lariat RNAs derived from the same intron or different lariat RNAs from the same pre-m RNA could be retained in one tissue while degraded in the other tissues.By deciphering lariat RNA sequences,we identified an AG-rich(RAAAAVAAAR)motif and a UC-rich(UCUCUYUCUC)motif for pollen-specific and the other three tissues-retained lariat RNAs,respectively.Reconstitution of the pollen-specific AG-rich motif indeed enhanced lariat RNA retention in plants.Biologically,hundreds of lariat RNAs harbored mi RNA binding sites,and dual-luciferase reporter assay showed that these natural lariat RNAs had the potential to protect expression of mi RNA target genes.Collectively,our results uncover that selective retention of lariat RNA is an actively regulatory process,and provide new insights into understanding how lariat RNA metabolism may impact mi RNA activity.

The chromatin remodeling complex imitation of switch controls stamen filament elongation by promoting jasmonic acid biosynthesis in Arabidopsis

Plant reproduction requires the coordinated development of both male and female reproductive organs.Jasmonic acid(JA)plays an essential role in stamen filament elongation.However,the mechanism by which the JA biosynthesis genes are regulated to promote stamen elongation remains unclear.Here,we show that the chromatin remodeling complex Imitation of Switch(ISWI)promotes stamen filament elongation by regulating JA biosynthesis.We show that AT-Rich Interacting Domain 5(ARID5)interacts with CHR11,CHR17,and RLT1,several known subunits of ISWI.Mutations in ARID5 and RLTs caused a reduced seed set due to greatly shortened stamen filaments.RNA-seq analyses reveal that the expression of key genes responsible for JA biosynthesis is significantly down-regulated in the arid5 and rlt mutants.Consistently,the JA levels are drastically decreased in both arid5 and rlt mutants.Chromatin immunoprecipitationquantitative PCR analyses further show that ARID5 is recruited to the chromatin of JA biosynthesis genes.Importantly,exogenous JA treatments can fully rescue the defects of stamen filament elongation in both arid5 and rlt mutants,leading to the partial recovery of fertility.Our results provide a clue how JA biosynthesisis positively regulated by the chromatin remodeling complex ISWI,thereby promoting stamen filament elongation in Arabidopsis.

A reciprocal inhibition between ARID1 and MET1 in male and female gametes in Arabidopsis

Both female and male gametophytes harbor companion cells and gametes. MET1, a DNA methyltransferase, is down-regulated in companion cells. However, how MET1 is differentially regulated in gametophytes remains unexplored. ARID1, a transcription factor that is specifically depleted in sperm cells, is occupied by MET1- dependent CG methylation. Here, we show that MET1 confines ARID1 to the vegetative cell of male gametes, but ARID1 conversely represses MET1 in the central cell of female gametes. Compared to the vegetative celllocalization in wild type pollen, ARID1 expands to sperm cells in the met1 mutant. To understand whether MET1- dependent ARID1 inhibition exists during female gametogenesis, we first show that ARID1 is expressed in the megaspore mother cell （MMC）, ARID1 but not MET1 is detectable in the central cell at maturity. Interestingly, compared to the absence of MET1 in the central cell and the egg cell of wild type ovules, MET1 significantly accumulates in these two cells in aridl ovules. Lastly, we show that both ARIDI and METI are required for the cell specification of MMC. Collectively, our results uncover a reciprocal dependence between ARIDI and METI, and provide a clue to further understand how the specification of MMC is likely regulated by DNA methylation.

A lariat-derived circular RNA is required for plant development in Arabidopsis

Lariat RNA is produced during pre-mRNA splicing, and it is traditionally thought as by-products, due to the quick turnover by debranching followed by degradation. However, recent findings identified many lariat RNAs accumulate with a circular form in higher eukaryotes. Although the remarkable accumulation, biological consequence of lariat-derived circular RNAs(here we name laciRNAs) remains largely unknown. Here, we report that a specific laciRNA from At5 g37720 plays an essential role in plant development by regulating gene expression globally. We focus on 17 laciRNAs with accumulation in wild type plants by circular RNA sequencing in Arabidopsis. To determine biological functions of these laciRNAs, we constructed one pair of transgenic plants for each laciRNA, in which the local gene with or without introns was over-expressed in wild type plants,respectively. By comparing morphological phenotypes and transcriptomic profiles between two classes of transgenic plants, we show that over-expression of the laciRNA derived from the 1 st intron of At5 g37720 causes pleiotropic phenotypes, including curly and clustered leaf, late flowering, reduced fertility, and accompanied with altered expression of approximately 800 genes.Our results provide another example that a specific plant circular RNA regulates gene expression in a similar manner to that of other non-coding RNAs under physiological conditions.

Cajal Bodies Are Developmentally Regulated during Pollen Development and Pollen Tube Growth in Arabidopsis thaliana

Dear Editor, Cajal bodies （CBs） are sub-nuclear bodies first described in neurons by Ramon y Cajal in 1903 and subsequently character- ized in both plant and animals （Gall, 2003）. They appear close to the nucleoli as discrete foci whose size and number vary during the cell cycle （Strzelecka et al., 2010）. They are dynamic structures that move, fuse, and separate, depending on the transcriptional status of the cell （Cioce and Lamond, 2005）. CBs are involved in assembly and trafficking of small nucleolar ribonucleopro- teins （snoRNPs） and in assembly and trafficking of spliceoso- mal small nuclear ribonucleoprotein （snRNPs） complexes that are involved in mRNA splicing （Gall, 2003; Cioce and Lamond, 2005）. CBs also contain components of the machinery involved in siRNA-mediated silencing and in methylation of repetitive DNA （Li et al., 2006）.

Brassinosteroids inhibit miRNA-mediated translational repression by decreasing AGO1 on the endoplasmic reticulum

Translational repression is a conserved mechanism in microRNA(miRNA)-guided gene silencing.In Arabidopsis,ARGONAUTE1(AGO1),the major miRNA effector,localizes in the cytoplasm for mRNA cleavage and at the endoplasmic reticulum(ER)for translational repression of target genes.However,the mechanism underlying miRNA-mediated translational repression is poorly understood.In particular,how the subcellular partitioning of AGO1 is regulated is largely unexplored.Here,we show that the plant hormone brassinosteroids(BRs)inhibit miRNA-mediated translational repression by negatively regulating the distribution of AGO1 at the ER in Arabidopsis thaliana.We show that the protein levels rather than the transcript levels of miRNA target genes were reduced in BR-deficient mutants but increased under BR treatments.The localization of AGO1 at the ER was significantly decreased under BR treatments while it was increased in the BR-deficient mutants.Moreover,ROTUNDIFOLIA3(ROT3),an enzyme involved in BR biosynthesis,co-localizes with AGO1 at the ER and interacts with AGO1 in a GW motif-dependent manner.Complementation analysis showed that the AGO1-ROT3 interaction is necessary for the function of ROT3.Our findings provide new clues to understand how miRNA-mediated gene silencing is regulated by plant endogenous hormones.