Biology subjects

Woodward, L. A.

Publications and source records attributed to Woodward, L. A..

3 recordsLinked to original sources

bioRxiv2025

PYM1 limits non-canonical Exon Junction Complex occupancy in a gene architecture dependent manner to tune mRNA expression

The Exon Junction Complex (EJC) deposited upstream of exon-exon junctions during pre-mRNA splicing in the nucleus remains stably bound to RNA to modulate mRNA fate at multiple post-transcriptional steps until its disassembly during translation. Here, we investigated two EJC disassembly mechanisms in human embryonic kidney 293 (HEK293) cells, one mediated by PYM1, a factor that can bind both the ribosome and the RBM8A/MAGOH heterodimer of the EJC core, and another by the elongating ribosome itself. We find that EJCs lacking PYM1 interaction show no defect in translation-dependent disassembly but is required for translation-independent EJC destabilization. Surprisingly, PYM1 interaction deficient EJCs are enriched on sites away from the canonical EJC binding position including on transcripts without introns or with fewer and longer exons. Acute reduction of PYM1 levels in HEK293 cells results in a modest inhibition of nonsense-mediated mRNA decay and stabilization of mRNAs that localize to endoplasmic reticulum associated TIS-granules and are characterized by fewer and longer exons. We confirmed the previously reported PYM1-flavivirus capsid protein interaction and found that human cells expressing the capsid protein or infected with flaviviruses show similar changes in gene expression as upon PYM1 depletion. Thus, PYM1 acts as an EJC specificity factor that is hijacked by flaviviruses to alter global EJC occupancy and reshape host cell mRNA regulation.

molecular biology↗

bioRxiv2024

Cycling DOF Factor mediated seasonal regulation of sexual reproduction and cold response is not conserved in Physcomitrium patens

Many land plants have evolved such that the transition from vegetative to reproductive development is synchronized with environmental cues. Examples of reproduction in response to seasonal cues can be found in both vascular and nonvascular species; however, most of our understanding of the molecular events controlling this timing has been worked out in angiosperm model systems. While the organism-level mechanisms of sexual reproduction vary dramatically between vascular and nonvascular plants, phylogenetic and transcriptomic evidence suggest paralogs in nonvascular plants may have conserved function with their vascular counterparts (Holm et al. 2010; Zhao et al. 2019; Genau et al. 2021). Given that Physcomitrium patens undergoes sexual reproductive development in response to photoperiodic and cold temperature cues (Hohe et al. 2002), it is well-suited for studying evolutionarily conserved mechanisms of seasonal control of reproduction. Thus, we used publicly available microarray data to identify genes differentially expressed in response to temperature cues (Fernandez-Pozo et al. 2020). We identified two CDF-like (CDL) genes in the P. patens genome that are the most like the angiosperm Arabidopsis thaliana CDFs based on conservation of protein motifs and diurnal expression patterns. In angiosperms, DNA-One Finger Transcription Factors (DOFs) play an important role in regulating photoperiodic flowering, regulating physiological changes in response to seasonal temperature changes, and mediating the cold stress response (Imaizumi et al. 2005; Kloosterman et al. 2013; Fornara et al. 2015; Ridge et al. 2016; Ding et al. 2018; Blair et al. 2022). We created knockout mutations and tested their impact on sexual reproduction and response to cold stress. Unexpectedly, the timing of sexual reproduction in the ppcdl double mutants did not differ significantly from wild type, suggesting that the PpCDLs are not necessary for seasonal regulation of this developmental transition. We also found that there was no change in expression of downstream cold-regulated genes in response to cold stress and no change in freezing tolerance in the knockout mutant plants. Finally, we observed no interaction between PpCDLs and the partial homologs of FKF1, an Arabidopsis thaliana repressor of CDFs. This is different from what is observed in angiosperms (Fornara et al. 2009; Corrales et al. 2014; Song et al. 2016, Li et al. 2013; Han et al. 2015, Wang et al. 2023, Li et al. 2009), which suggests that the functions of CDF proteins in angiosperms are not conserved in P. patens.

plant biology↗

bioRxiv2018

The exon junction complex undergoes a compositional switch that alters mRNP structure and nonsense-mediated mRNA decay activity

The exon junction complex (EJC) deposited upstream of mRNA exon junctions shapes structure, composition and fate of spliced mRNA ribonucleoprotein particles (mRNPs). To achieve this, the EJC core nucleates assembly of a dynamic shell of peripheral proteins that function in diverse post-transcriptional processes. To illuminate consequences of EJC composition change, we purified EJCs from human cells via peripheral proteins RNPS1 and CASC3. We show that EJC originates as an SR-rich mega-dalton sized RNP that contains RNPS1 but lacks CASC3. After mRNP export to the cytoplasm and before translation, the EJC undergoes a remarkable compositional and structural remodeling into an SR-devoid monomeric complex that contains CASC3. Surprisingly, RNPS1 is important for nonsense-mediated mRNA decay (NMD) in general whereas CASC3 is needed for NMD of only select mRNAs. The promotion of switch to CASC3-EJC slows down NMD. Overall, the EJC compositional switch dramatically alters mRNP structure and specifies two distinct phases of EJC-dependent NMD.

molecular biology↗