IRE1α Mediates the Hypertrophic Growth of Cardiomyocytes Through Facilitating the Formation of Initiation Complex to Promote the Translation of TOP-Motif Transcripts.

BACKGROUND: Cardiomyocyte growth is coupled with active protein synthesis, which is one of the basic biological processes in living cells. However, it is unclear whether the unfolded protein response transducers and effectors directly take part in the control of protein synthesis. The connection between critical functions of the unfolded protein response in cellular physiology and requirements of multiple processes for cell growth prompted us to investigate the role of the unfolded protein response in cell growth and underlying molecular mechanisms. METHODS: Cardiomyocyte-specific inositol-requiring enzyme 1α (IRE1α) knockout and overexpression mouse models were generated to explore its function in vivo. Neonatal rat ventricular myocytes were isolated and cultured to evaluate the role of IRE1α in cardiomyocyte growth in vitro. Mass spectrometry was conducted to identify novel interacting proteins of IRE1α. Ribosome sequencing and polysome profiling were performed to determine the molecular basis for the function of IRE1α in translational control. RESULTS: We show that IRE1α is required for cell growth in neonatal rat ventricular myocytes under prohypertrophy treatment and in HEK293 cells in response to serum stimulation. At the molecular level, IRE1α directly interacts with eIF4G and eIF3, 2 critical components of the translation initiation complex. We demonstrate that IRE1α facilitates the formation of the translation initiation complex around the endoplasmic reticulum and preferentially initiates the translation of transcripts with 5' terminal oligopyrimidine motifs. We then reveal that IRE1α plays an important role in determining the selectivity and translation of these transcripts. We next show that IRE1α stimulates the translation of epidermal growth factor receptor through an unannotated terminal oligopyrimidine motif in its 5' untranslated region. We further demonstrate a physiological role of IRE1α-governed protein translation by showing that IRE1α is essential for cardiomyocyte growth and cardiac functional maintenance under hemodynamic stress in vivo. CONCLUSIONS: These studies suggest a noncanonical, essential role of IRE1α in orchestrating protein synthesis, which may have important implications in cardiac hypertrophy in response to pressure overload and general cell growth under other physiological and pathological conditions.

Prostaglandins limit nuclear actin to control nucleolar function during oogenesis.

Prostaglandins (PGs), locally acting lipid signals, regulate female reproduction, including oocyte development. However, the cellular mechanisms of PG action remain largely unknown. One cellular target of PG signaling is the nucleolus. Indeed, across organisms, loss of PGs results in misshapen nucleoli, and changes in nucleolar morphology are indicative of altered nucleolar function. A key role of the nucleolus is to transcribe ribosomal RNA (rRNA) to drive ribosomal biogenesis. Here we take advantage of the robust, in vivo system of Drosophila oogenesis to define the roles and downstream mechanisms whereby PGs regulate the nucleolus. We find that the altered nucleolar morphology due to PG loss is not due to reduced rRNA transcription. Instead, loss of PGs results in increased rRNA transcription and overall protein translation. PGs modulate these nucleolar functions by tightly regulating nuclear actin, which is enriched in the nucleolus. Specifically, we find that loss of PGs results in both increased nucleolar actin and changes in its form. Increasing nuclear actin, by either genetic loss of PG signaling or overexpression of nuclear targeted actin (NLS-actin), results in a round nucleolar morphology. Further, loss of PGs, overexpression of NLS-actin or loss of Exportin 6, all manipulations that increase nuclear actin levels, results in increased RNAPI-dependent transcription. Together these data reveal PGs carefully balance the level and forms of nuclear actin to control the level of nucleolar activity required for producing fertilization competent oocytes.

Importin-9 regulates chromosome segregation and packaging in Drosophila germ cells.

Germ cells undergo distinct nuclear processes as they differentiate into gametes. Although these events must be coordinated to ensure proper maturation, the stage-specific transport of proteins in and out of germ cell nuclei remains incompletely understood. Our efforts to genetically characterize Drosophila genes that exhibit enriched expression in germ cells led to the finding that loss of the highly conserved Importin ß/karyopherin family member Importin-9 (Ipo9, herein referring to Ranbp9) results in female and male sterility. Immunofluorescence and fluorescent in situ hybridization revealed that Ipo9KO mutants display chromosome condensation and segregation defects during meiosis. In addition, Ipo9KO mutant males form abnormally structured sperm and fail to properly exchange histones for protamines. Ipo9 physically interacts with proteasome proteins, and Ipo9 mutant males exhibit disruption of the nuclear localization of several proteasome components. Thus, Ipo9 coordinates the nuclear import of functionally related factors necessary for the completion of gametogenesis. This article has an associated First Person interview with the first author of the paper.

Functions for Cdc42p BEM adaptors in regulating a differentiation-type MAP kinase pathway.

Ras homology (Rho) GTPases regulate cell polarity and signal transduction pathways to control morphogenetic responses in different settings. In yeast, the Rho GTPase Cdc42p regulates cell polarity, and through the p21-activated kinase Ste20p, Cdc42p also regulates mitogen-activated protein kinase (MAPK) pathways (mating, filamentous growth or fMAPK, and HOG). Although much is known about how Cdc42p regulates cell polarity and the mating pathway, how Cdc42p regulates the fMAPK pathway is not clear. To address this question, Cdc42p-dependent MAPK pathways were compared in the filamentous (Σ1278b) strain background. Each MAPK pathway showed a unique activation profile, with the fMAPK pathway exhibiting slow activation kinetics compared with the mating and HOG pathways. A previously characterized version of Cdc42p, Cdc42pE100A, that is specifically defective for fMAPK pathway signaling, was defective for interaction with Bem4p, the pathway-specific adaptor for the fMAPK pathway. Corresponding residues in Bem4p were identified that were required for interaction with Cdc42p and fMAPK pathway signaling. The polarity adaptor Bem1p also regulated the fMAPK pathway. Versions of Bem1p defective for recruitment of Ste20p to the plasma membrane, intramolecular interactions, and interaction with the GEF, Cdc24p, were defective for fMAPK pathway signaling. Bem1p also regulated effector pathways in different ways. In some pathways, multiple domains of the protein were required for its function, whereas in other pathways, a single domain or function was needed. Genetic suppression tests showed that Bem4p and Bem1p regulate the fMAPK pathway in an ordered sequence. Collectively, the study demonstrates unique and sequential functions for Rho GTPase adaptors in regulating MAPK pathways.

Partial Inhibition of mTORC1 in Aged Rats Counteracts the Decline in Muscle Mass and Reverses Molecular Signaling Associated with Sarcopenia.

There is a lack of pharmacological interventions available for sarcopenia, a progressive age-associated loss of muscle mass, leading to a decline in mobility and quality of life. We found mTORC1 (mammalian target of rapamycin complex 1), a well-established positive modulator of muscle mass, to be surprisingly hyperactivated in sarcopenic muscle. Furthermore, partial inhibition of the mTORC1 pathway counteracted sarcopenia, as determined by observing an increase in muscle mass and fiber type cross-sectional area in select muscle groups, again surprising because mTORC1 signaling has been shown to be required for skeletal muscle mass gains in some models of hypertrophy. Additionally, several genes related to senescence were downregulated and gene expression indicators of neuromuscular junction denervation were diminished using a low dose of a "rapalog" (a pharmacological agent related to rapamycin). Therefore, partial mTORC1 inhibition may delay the progression of sarcopenia by directly and indirectly modulating multiple age-associated pathways, implicating mTORC1 as a therapeutic target to treat sarcopenia.