Chronic kidney disease alters Pin1 phosphorylation and parathyroid hormone mRNA binding proteins leading to secondary hyperparathyroidism

Parathyroid hormone (PTH) regulates calcium metabolism and bone strength. Chronic kidney disease (CKD) leads to secondary hyperparathyroidism (SHP) which increases morbidity and mortality. High PTH expression in SHP is due to increased PTH mRNA stability mediated by changes in PTH mRNA interaction with stabilizing AUF1 and destabilizing KSRP. Pin1 isomerizes target proteins, including mRNA binding proteins. In SHP, Pin1 isomerase activity is decreased and phosphorylated KSRP fails to bind PTH mRNA, resulting in high PTH mRNA stability and levels. The molecular mechanisms underlying Pin1 regulation and their effect to increase PTH expression are unknown. We show by mass-spectrometry (MS) the CKD induced changes in rat parathyroid proteome and phosphoproteome profiles. Parathyroid Pin1 Ser16 and Ser71 phosphorylation, that disrupts Pin1 activity, is enhanced in acute and chronic kidney failure rats. Accordingly, pharmacologic Pin1 inhibition increases PTH expression in parathyroid organ cultures and transfected cells, through the PTH mRNA protein binding cis element and KSRP phosphorylation. Therefore, CKD leads to parathyroid loss of Pin1 activity by inducing Pin1 phosphorylation. This predisposes parathyroids to increase PTH production through modified PTH mRNA-KSRP interaction that is dependent on KSRP phosphorylation. CKD induced Pin1 and KSRP phosphorylation and the Pin1-KSRP-PTH mRNA axis thus drive secondary hyperparathyroidism.

The endoplasmic reticulum-associated mRNA-binding proteins ERBP1 and ERBP2 interact in bloodstream-form Trypanosoma brucei

Kinetoplastids rely heavily on post-transcriptional mechanisms for control of gene expression, and on RNA-binding proteins that regulate mRNA splicing, translation and decay. Trypanosoma brucei ERBP1 (Tb927.10.14150) and ERBP2 (Tb927.9.9550) were previously identified as mRNA binding proteins that lack canonical RNA-binding domains. We show here that ERBP1 is associated with the endoplasmic reticulum, like ERBP2, and that the two proteins interact in vivo. Loss of ERBP1 from bloodstream-form T. brucei initially resulted in a growth defect but proliferation was restored after more prolonged cultivation. Pull-down analysis of tagged ERBP1 suggests that it preferentially binds to ribosomal protein mRNAs. The ERBP1 sequence resembles that of Saccharomyces cerevisiae Bfr1, which also localises to the endoplasmic reticulum and binds to ribosomal protein mRNAs. However, unlike Bfr1, ERBP1 does not bind to mRNAs encoding secreted proteins, and it is also not recruited to stress granules after starvation.

yRACK1/Asc1 proxiOMICs—Towards Illuminating Ships Passing in the Night

Diverse signals and stress factors regulate the activity and homeostasis of ribosomes in all cells. The Saccharomyces cerevisiae protein Asc1/yRACK1 occupies an exposed site at the head region of the 40S ribosomal subunit (hr40S) and represents a central hub for signaling pathways. Asc1 strongly affects protein phosphorylation and is involved in quality control pathways induced by translation elongation arrest. Therefore, it is important to understand the dynamics of protein formations in the Asc1 microenvironment at the hr40S. We made use of the in vivo protein-proximity labeling technique Biotin IDentification (BioID). Unbiased proxiOMICs from two adjacent perspectives identified nucleocytoplasmic shuttling mRNA-binding proteins, the deubiquitinase complex Ubp3-Bre5, as well as the ubiquitin E3 ligase Hel2 as neighbors of Asc1. We observed Asc1-dependency of hr40S localization of mRNA-binding proteins and the Ubp3 co-factor Bre5. Hel2 and Ubp3-Bre5 are described to balance the mono-ubiquitination of Rps3 (uS3) during ribosome quality control. Here, we show that the absence of Asc1 resulted in massive exposure and accessibility of the C-terminal tail of its ribosomal neighbor Rps3 (uS3). Asc1 and some of its direct neighbors together might form a ribosomal decision tree that is tightly connected to close-by signaling modules.

The endoplasmic reticulum-associated mRNA-binding proteins ERBP1 and ERBP2 interact in bloodstream-form Trypanosoma brucei

Kinetoplastids rely heavily on post-transcriptional mechanisms for control of gene expression, and on RNA-binding proteins that regulate mRNA splicing, translation and decay. Trypanosoma brucei ERBP1 (Tb927.10.14150) and ERBP2 (Tb927.9.9550) were previously identified as mRNA binding proteins that lack canonical RNA-binding domains. We here show that ERBP1 is associated with the endoplasmic reticulum, like ERBP2, and that the two proteins interact in vivo. Loss of ERBP1 from bloodstream-form T. brucei initially resulted in a growth defect but proliferation was restored after more prolonged cultivation. Results from a pull-down of tagged ERBP1 suggest that it preferentially binds to ribosomal protein mRNAs. The ERBP1 sequence resembles that of Saccharomyces cerevisiae Bfr1, which also localises to the endoplasmic reticulum and binds to ribosomal protein mRNAs. However, unlike Bfr1, ERBP1 does not bind to mRNAs encoding secreted proteins, and it is also not recruited to stress granules after starvation.

Circadian clock-dependent and -independent posttranscriptional regulation underlies temporal mRNA accumulation in mouse liver

The mammalian circadian clock coordinates physiology with environmental cycles through the regulation of daily oscillations of gene expression. Thousands of transcripts exhibit rhythmic accumulations across mouse tissues, as determined by the balance of their synthesis and degradation. While diurnally rhythmic transcription regulation is well studied and often thought to be the main factor generating rhythmic mRNA accumulation, the extent of rhythmic posttranscriptional regulation is debated, and the kinetic parameters (e.g., half-lives), as well as the underlying regulators (e.g., mRNA-binding proteins) are relatively unexplored. Here, we developed a quantitative model for cyclic accumulations of pre-mRNA and mRNA from total RNA-seq data, and applied it to mouse liver. This allowed us to identify that about 20% of mRNA rhythms were driven by rhythmic mRNA degradation, and another 15% of mRNAs regulated by both rhythmic transcription and mRNA degradation. The method could also estimate mRNA half-lives and processing times in intact mouse liver. We then showed that, depending on mRNA half-life, rhythmic mRNA degradation can either amplify or tune phases of mRNA rhythms. By comparing mRNA rhythms in wild-type and Bmal1−/− animals, we found that the rhythmic degradation of many transcripts did not depend on a functional BMAL1. Interestingly clock-dependent and -independent degradation rhythms peaked at distinct times of day. We further predicted mRNA-binding proteins (mRBPs) that were implicated in the posttranscriptional regulation of mRNAs, either through stabilizing or destabilizing activities. Together, our results demonstrate how posttranscriptional regulation temporally shapes rhythmic mRNA accumulation in mouse liver.