The Arabidopsis THO/TREX component TEX1 functionally interacts with MOS11 and modulates mRNA export and alternative splicing events

Sørensen, Brian B.; Ehrnsberger, Hans F.; Susan, Esposito; Pfab, Alexander; Bruckmann, Astrid; Hauptmann, Judith; Meister, Gunter; Merkl, Rainer; Schubert, Thomas; Längst, Gernot; Melzer, Michael; Grasser, Marion; Grasser, Klaus D.

doi:10.1007/s11103-016-0561-9

Cited by 36 publications

(61 citation statements)

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“…The core complex of Arabidopsis THO/TREX consists of HPR1 (THO1), THO2, THO5A/B, THO6, THO7A/B and TEX1 (66) associated with UAP56, MOS11 and the mRNA export adapters THO4B-C (aka ALY2-4) (67). Using TEX1, UAP56 and MOS11 as baits, Arabidopsis THO/TREX was shown to associate with a number of splicing factors and with components of the nuclear pore complex (67). Our data now extend these observations by showing that the association of THO/TREX with SE/ARS2 is also conserved in plants.…”

Section: Discussionmentioning

confidence: 99%

SERRATE interacts with the nuclear exosome targeting (NEXT) complex to degrade primary miRNA precursors in Arabidopsis

Bajczyk

Lange

Bielewicz

et al. 2020

Preprint

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SERRATE/ARS2 is a conserved RNA effector protein involved in transcription, processing and export of different types of RNAs. In Arabidopsis, the best-studied function of SERRATE (SE) is to promote miRNA processing. Here, we report that SE interacts with the Nuclear Exosome Targeting (NEXT) complex, comprising the RNA helicase HEN2, the RNA binding protein RBM7 and one of the two zincknuckle proteins ZCCHC8A/ZCCHC8B. The identification of common targets of SE and HEN2 by RNAseq supports the idea that SE cooperates with NEXT for RNA surveillance by the nuclear exosome.Among the RNA targets accumulating in absence of SE or NEXT are miRNA precursors. Loss of NEXT components results in the accumulation of pri-miRNAs without affecting levels of miRNAs, indicating that NEXT is, unlike SE, not required for miRNA processing. As compared to se-2, se-2 hen2-2 double mutants showed increased accumulation of pri-miRNAs, but partially restored levels of mature miRNAs and attenuated developmental defects. We propose that the slow degradation of pri-miRNAs caused by loss of HEN2 compensates for the poor miRNA processing efficiency in se-2 mutants, and that SE regulates miRNA biogenesis through its double contribution in promoting miRNA processing but also pri-miRNA degradation through the recruitment of the NEXT complex. IntroductionIn plants, many miRNAs are encoded by independent RNA polymerase II transcription units. The primary transcripts contain a 5' cap structure as well as a poly(A) tail at the 3' end (1), and sometimes introns. The primary or spliced pri-miRNAs adopt stem loop structures which are processed by the nuclear endonuclease Dicer like 1 (DCL1) (2) in two sequential reactions. The first step creates shorter miRNA precursors called pre-miRNAs. The second step produces miRNA/miRNA* duplexes of mostly 21 nt with 2 nt overhangs at both 3' ends. DCL1 associates with the double stranded RNA binding 2 protein HYPONASTIC LEAVES 1 (HYL1) and the zinc finger domain-containing RNA effector protein SERRATE (3-5). Both HYL1 and SE interact with DCL1 as well as with each other (6-8). DCL1, SE and HYL1 form the core of the plant Microprocessor complex and co-localize within the nucleus in special structures known as dicing bodies (D-bodies) (7). In absence of HYL1 or upon reduced activity of SE, dicing by DCL1 is not completely abolished but less efficient and less accurate (3-5). The importance of SE for the biogenesis of miRNAs is illustrated by the phenotype of se mutants. Complete loss of SE expression as in the null mutant se-4 is embryonic lethal, while the se-1 mutation, which results in the expression of a truncated SE lacking 20 amino acids at its C-terminus, severely affects developmental timing, phylotaxy, meristem function and patterning of leaves and flowers (4,9). The se-2 mutation, in which the SE protein is truncated by 40 amino acids from the C-terminus, additionally displays the hyponastic leave shape that is characteristic for many miRNA biogenesis mutants including hyl1-2, hst-1 and ago1-25 (10-13).In ad...

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Section: Discussionmentioning

confidence: 99%

SERRATE interacts with the nuclear exosome targeting (NEXT) complex to degrade primary miRNA precursors in Arabidopsis

Bajczyk

Lange

Bielewicz

et al. 2020

Preprint

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“…Arabidopsis cells (Sørensen et al 2017) and ALY2 interacts with the RNA helicase UAP56 in 183 yeast cells and in vitro (Kammel et al 2013 analyses (Kammel et al 2013), UAP56-GFP was detected exclusively in the cell nuclei ( Fig. …”

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confidence: 97%

“…ALYs and MOS11 (the orthologue of CIP29) in Arabidopsis cells (Sørensen et al 2017 (Germain et al 2010;Lu et al 2010;Pan et al 2012;Xu et al 2015; 114 Sørensen et al 2017), but so far none of the ALY mRNA export adaptor candidates have been…”

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ALY RNA-Binding Proteins Are Required for Nucleocytosolic mRNA Transport and Modulate Plant Growth and Development

et al. 2018

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3 ABSTRACT 49The regulated transport of mRNAs from the cell nucleus to the cytosol is a critical step 50 linking transcript synthesis and processing with translation. However, in plants, only few of 51 the factors that act in the mRNA export pathway have been functionally characterised. 52Flowering plant genomes encode several members of the ALY protein family, which function 53 as mRNA export factors in other organisms. Arabidopsis thaliana ALY1-4 are commonly 54 detected in root and leaf cells, but are differentially expressed in reproductive tissue. 55Moreover, the subnuclear distribution of ALY1/2 differs from that of ALY3/4. ALY1 binds 56 with higher affinity to ssRNA than dsRNA and ssDNA, and interacts preferentially with 5- THO rather than that of yeast (Yelina et al. 2010). Likewise, THO associates with UAP56, 99ALYs and MOS11 (the orthologue of CIP29) in Arabidopsis cells (Sørensen et al. 2017 (Germain et al. 2010;Lu et al. 2010;Pan et al. 2012;Xu et al. 2015; 114 Sørensen et al. 2017), but so far none of the ALY mRNA export adaptor candidates have been 115shown to function in nucleo-cytosolic transport of mRNAs in plants. 116In this study, we have systematically studied the Arabidopsis ALY proteins including their RESULTS 126 ALY proteins in Arabidopsis and other plants 127First, we compared the amino acid sequences of the four Arabidopsis ALY proteins (ALY1- sequence identity) as well as ALY3 and ALY4 (70% amino acid sequence identity) share a 137 high degree of sequence similarity, whereas the similarity of ALY1/2 versus ALY3/4 is 138 clearly lower (<42% amino acid sequence identity) (Supplemental Fig. S1B (Fig. 1A). ALY1 and truncated versions of the protein were expressed in 157 E. coli as 6xHis-GB1 fusion proteins, purified by two-step chromatography, and examined by 158 SDS-PAGE (Fig. 1B). For comparison we also used the unfused 6xHis-GB1 tag. The purified (Fig. 1C, Student's t-test, P < 0.05 and P < 0.001, respectively). The unfused 6xHis-GB1 165 tag did not exhibit affinity for ssRNA. To examine which domains of ALY1 contribute to the 166 RNA interactions, the binding to ssRNA of the different recombinant ALY1 versions was 167 measured (Fig. 1D (Fig. 3C, D). Therefore, the four ALY proteins and UAP56 228 are widely expressed in sporophytic cells of Arabidopsis plants. 229The subnuclear localisation of the ALY-GFP fusions was inspected in more detail by 230 CLSM in comparison to the UAP56-GFP and GFP-NLS controls. GFP-NLS (Antosch et al. immunofluorescence microscopy analysis (Kammel et al. 2013 (Table S1). In leaf cells, the nucleoplasmic distribution of 238 the ALY proteins appeared more heterogeneous than in root cells particularly for ALY4 that 239 partially localised to nucleoplasmic foci (Fig. 4B). Notably, the nucleolar enrichment of 253In view of the apparently ubiquitous expression of the four ALY-GFP proteins in 254 sporophytic cells, their occurrence was analysed in male and female gametophytes by CLSM. 255In mature pollen grains (Fig. 5A), the fluorescent signal of ALY1-...

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“…Previous studies implicated the THOC in multiple steps of mRNA maturation, mRNP formation and nuclear export (Luna et al, 2012; Sørensen et al, 2017; Tran et al, 2014a; Wang et al, 2013). In order to elucidate THOC’s role in presynaptic gene expression we performed single molecule in-situ hybridization (smFISH) experiments to specifically visualize and determine the intracellular localization of presynaptic as well as control transcripts in PDE DA neurons.…”

Section: Resultsmentioning

confidence: 99%

A conserved nuclear export complex coordinates transcripts for dopaminergic synaptogenesis and neuronal surviva

Kaganovsky

et al. 2018

Preprint

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Summary Synaptic vesicle and active zone proteins are required for synaptogenesis. The molecular mechanisms for coordinated synthesis of these proteins are not understood. Using forward genetic screens, we identified the conserved THO nuclear export Complex (THOC) as master regulator of presynapse development in C.elegans dopaminergic neurons. In THOC mutants, synaptic messenger RNAs are trapped in the nucleus, resulting in dramatic decrease of synaptic protein expression, near complete loss of synapses and compromised dopamine function. cAMP-responsive element binding protein (CREB) interacts with THOC to mark activity-dependent transcripts for efficient nuclear export. Deletion of the THOC subunit Thoc5 in mouse dopaminergic neurons causes severe defects in synapse maintenance and subsequent neuronal death in the Substantia Nigra compacta (SNc). These cellular defects lead to abrogated dopamine release, ataxia and animal death. Together, our results argue that nuclear export mechanisms can select specific mRNAs and be a rate-limiting step for synapse development and neuronal survival. Highlights Dopaminergic presynapses are severely impaired in thoc mutant worms and mice THOC specifically controls the nuclear export of synaptic transcripts CREB recruits THOC onto activity-dependent synaptic transcripts for efficient export Dopamine neurons in the SNc degenerate upon conditional knock-out of thoc5

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The Arabidopsis THO/TREX component TEX1 functionally interacts with MOS11 and modulates mRNA export and alternative splicing events

Cited by 36 publications

References 66 publications

SERRATE interacts with the nuclear exosome targeting (NEXT) complex to degrade primary miRNA precursors in Arabidopsis

SERRATE interacts with the nuclear exosome targeting (NEXT) complex to degrade primary miRNA precursors in Arabidopsis

ALY RNA-Binding Proteins Are Required for Nucleocytosolic mRNA Transport and Modulate Plant Growth and Development

A conserved nuclear export complex coordinates transcripts for dopaminergic synaptogenesis and neuronal surviva

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