The mechanisms by which poikilothermic organisms ensure that biological processes are robust to temperature changes are largely unknown. Temperature compensation, the ability of circadian rhythms to maintain a relatively constant period over the broad range of temperatures resulting from seasonal fluctuations in environmental conditions, is a defining property of circadian networks. Temperature affects the alternative splicing (AS) of several clock genes in fungi, plants, and flies, but the splicing factors that modulate these effects to ensure clock accuracy throughout the year remain to be identified. Here we show that GEMIN2, a spliceosomal small nuclear ribonucleoprotein assembly factor conserved from yeast to humans, modulates low temperature effects on a large subset of pre-mRNA splicing events. In particular, GEMIN2 controls the AS of several clock genes and attenuates the effects of temperature on the circadian period in Arabidopsis thaliana. We conclude that GEMIN2 is a key component of a posttranscriptional regulatory mechanism that ensures the appropriate acclimation of plants to daily and seasonal changes in temperature conditions. Significance RNA processing, an important step in the regulation of gene expression, is mediated by proteins and RNA molecules that are highly sensitive to variations in temperature conditions. Most organisms do not control their own body temperature. Therefore, molecular mechanisms must have evolved that ensure that biological processes are robust to temperature changes. Here we identify a protein that buffers the effect of temperature on biological timing by enhancing the assembly of the spliceosome, a large ribonucleoprotein complex involved in RNA processing in organisms ranging from yeast to humans, and thereby controlling the alternative splicing of clock genes.

Significance: There is increasing evidence that previously considered core constituents of multi-subunit complexes involved in RNA processing play regulatory rather than passive roles in the control of gene expression, but specific signaling pathways in which they participate are not known. Here we show that SM-like ( LSM ) genes, which encode core components of the spliceosome, are regulated by the circadian clock and control clock function in plants and mammals, revealing convergent evolutionary mechanisms mediating posttranscriptional regulation of circadian networks across kingdoms.

Arabidopsis (Arabidopsis thaliana) PROTEIN ARGININE METHYLTRANSFERASE5 (PRMT5) post-translationally modifies RNA-binding proteins by arginine (R) methylation. However, the impact of this modification on the regulation of RNA processing is largely unknown. We used the spliceosome component, SM-LIKE PROTEIN 4 (LSM4), as a paradigm to study the role of R-methylation in RNA processing. We found that LSM4 regulates alternative splicing (AS) of a suite of its in vivo targets identified here. The lsm4 and prmt5 mutants show a considerable overlap of genes with altered AS raising the possibility that splicing of those genes could be regulated by PRMT5-dependent LSM4 methylation. Indeed, LSM4 methylation impacts AS, particularly of genes linked with stress response. Wild-type LSM4 and an unmethylable version complement the lsm4-1 mutant, suggesting that methylation is not critical for growth in normal environments. However, LSM4 methylation increases with abscisic acid and is necessary for plants to grow under abiotic stress. Conversely, bacterial infection reduces LSM4 methylation, and plants that express unmethylable-LSM4 are more resistant to Pseudomonas than those expressing wild-type LSM4. This tolerance correlates with decreased intron retention of immune-response genes upon infection. Taken together, this provides direct evidence that R-methylation adjusts LSM4 function on pre-mRNA splicing in an antagonistic manner in response to biotic and abiotic stress.

Circadian clocks, the molecular devices present in almost all eukaryotic and some prokaryotic organisms, phase biological activities to the most appropriate time of day. These devices are synchronized by the daily cycles of light and temperature, and control hundreds of processes, ranging from gene expression to behavior as well as reproductive development. For a long time, these clocks were considered to operate primarily through regulatory feedback loops that act at the transcriptional level. Recent studies, however, conclusively show that circadian rhythms can persist in the absence of transcription, and it is evident that robust and precise circadian oscillations require multiple regulatory mechanisms operating at the co-/post-transcriptional, translational, post-translational and metabolic levels. Furthermore, these different regulatory loops exhibit strong interactions, which contribute to the synchronization of biological rhythms with environmental changes throughout the day and year. Here, we describe recent advances that highlight the role of alternative splicing (AS) in the operation of circadian networks, focusing on molecular and genomic studies conducted in Arabidopsis thaliana. These studies have also enhanced our understanding of the mechanisms that control AS and of the physiological impact of AS.