The cytoplasmic polyadenylation element-binding (CPEB) proteins regulate pre-mRNA processing and translation of CPE-containing mRNAs in early embryonic development and synaptic activity. However, specific functions in adult organisms are poorly understood. Here we show that CPEB4 is required for adaptation to high-fat-diet- and ageing-induced endoplasmic reticulum (ER) stress, and subsequent hepatosteatosis. Stress-activated liver CPEB4 expression is dual-mode regulated. First, Cpeb4 mRNA transcription is controlled by the circadian clock, and then its translation is regulated by the unfolded protein response (UPR) through upstream open reading frames within the 5'UTR. Thus, the CPEB4 protein is synthesized only following ER stress but the induction amplitude is circadian. In turn, CPEB4 activates a second wave of UPR translation required to maintain ER and mitochondrial homeostasis. Our results suggest that combined transcriptional and translational Cpeb4 regulation generates a 'circadian mediator', which coordinates hepatic UPR activity with periods of high ER-protein-folding demand. Accordingly, CPEB4 deficiency results in non-alcoholic fatty liver disease.

RNA-binding proteins (RBPs) control critical aspects of cardiomyocyte function, but the repertoire of active RBPs in cardiomyocytes during the growth response is largely unknown. We define RBPs in healthy and diseased cardiomyocytes at a system-wide level by RNA interactome capture. This identifies 67 cardiomyocyte-specific RBPs, including several contractile proteins. Furthermore, we identify the cytoplasmic polyadenylation element-binding protein 4 (Cpeb4) as a dynamic RBP, regulating cardiac growth both in vitro and in vivo. We identify mRNAs bound to and regulated by Cpeb4 in cardiomyocytes. Cpeb4 regulates cardiac remodeling by differential expression of transcription factors. Among Cpeb4 target mRNAs, two zinc finger transcription factors (Zeb1 and Zbtb20) are discovered. We show that Cpeb4 regulates the expression of these mRNAs and that Cpeb4 depletion increases their expression. Thus, Cpeb4 emerges as a critical regulator of cardiomyocyte function by differential binding to specific mRNAs in response to pathological growth stimulation.

Organogenesis is directed by coordinated cell proliferation and differentiation programs. The hierarchical networks of transcription factors driving mammary gland development and function have been widely studied. However, the contribution of posttranscriptional gene expression reprogramming remains largely unexplored. The 3′ untranslated regions of messenger RNAs (mRNAs) contain combinatorial ensembles of cis-regulatory elements that define transcript-specific regulation of protein synthesis through their cognate RNA binding proteins. We analyze the contribution of the RNA binding cytoplasmic polyadenylation element–binding (CPEB) protein family, which collectively regulate mRNA translation for about 30% of the genome. We find that CPEB2 is required for the integration of hormonal signaling by controlling the protein expression from a subset of ER/PR- regulated transcripts. Furthermore, CPEB2 is critical for the development of ER-positive breast tumors. This work uncovers a previously unknown gene expression regulation level in breast morphogenesis and tumorigenesis, coordinating sequential transcriptional and posttranscriptional layers of gene expression regulation.

Chronic inflammation is a major cause of disease. Inflammation resolution is in part directed by the differential stability of mRNAs encoding pro-inflammatory and anti-inflammatory factors. In particular, tristetraprolin (TTP)-directed mRNA deadenylation destabilizes AU-rich element (ARE)-containing mRNAs. However, this mechanism alone cannot explain the variety of mRNA expression kinetics that are required to uncouple degradation of pro-inflammatory mRNAs from the sustained expression of anti-inflammatory mRNAs. Here we show that the RNA-binding protein CPEB4 acts in an opposing manner to TTP in macrophages: it helps to stabilize anti-inflammatory transcripts harboring cytoplasmic polyadenylation elements (CPEs) and AREs in their 3′-UTRs, and it is required for the resolution of the LPS-triggered inflammatory response. Coordination of CPEB4 and TTP activities is sequentially regulated through MAPK signaling. Accordingly, CPEB4 depletion in macrophages impairs inflammation resolution in an LPS-induced sepsis model. We propose that the counterbalancing actions of CPEB4 and TTP, as well as the distribution of CPEs and AREs in their target mRNAs, define transcript-specific decay patterns required for inflammation resolution. Thus, these two opposing mechanisms provide a fine-tuning control of inflammatory transcript destabilization while maintaining the expression of the negative feedback loops required for efficient inflammation resolution; disruption of this balance can lead to disease.

Tumor growth is influenced by a complex network of interactions between multiple cell types in the tumor microenvironment (TME). These constrained conditions trigger the endoplasmic reticulum (ER) stress response, which extensively reprograms mRNA translation. When uncontrolled over time, chronic ER stress impairs the antitumor effector function of CD8 T lymphocytes. How cells promote adaptation to chronic stress in the TME without the detrimental effects of the terminal unfolded protein response (UPR) is unknown. Here, we find that, in effector CD8 T lymphocytes, RNA‐binding protein CPEB4 constitutes a new branch of the UPR that allows cells to adapt to sustained ER stress, yet remains decoupled from the terminal UPR. ER stress, induced during CD8 T‐cell activation and effector function, triggers CPEB4 expression. CPEB4 then mediates chronic stress adaptation to maintain cellular fitness, allowing effector molecule production and cytotoxic activity. Accordingly, this branch of the UPR is required for the antitumor effector function of T lymphocytes, and its disruption in these cells exacerbates tumor growth.

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The inclusion of microexons by alternative splicing is frequent in neuronal proteins. The roles of these sequences are in most cases unknown, but changes in their degree of inclusion are associated with neurodevelopmental diseases. We recently found that the decreased inclusion of a 24-nucleotide neuron-specific microexon in CPEB4, an RNA-binding protein that regulates translation through cytoplasmic changes in poly(A) tail length, is linked to idiopathic autism spectrum disorder (ASD). Why this microexon is required and how small changes in its degree of inclusion generate a dominant-negative effect on the expression of ASD-linked genes is not clear. Here we show that neuronal CPEB4 forms condensates that dissolve upon depolarization, a phase transition associated with a switch from translational repression to activation. Heterotypic intermolecular interactions between the microexon and a cluster of histidine residues kinetically stabilize the condensates by competing with homotypic interactions between clusters, that otherwise lead to the irreversible aggregation of CPEB4. We conclude that microexon 4 in neuronal CPEB4 is required to preserve the reversible regulation of CPEB4-mediated gene expression in response to neuronal stimulation.

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