Autophagy is a tightly regulated catabolic process wherein cells under stress sequester cytosolic constituents like damaged proteins and organelles in double-membrane vesicles called autophagosomes. The autophagosomes degrade their cargo by lysosomal proteolysis generating raw materials for the biosynthesis of vital macromolecules. One of the initial steps in the assembly of autophagosomes from pre-autophagic structures is the recruitment and activation of the class III phosphatidylinositol 3-kinase complex consisting of Beclin 1 (BECN1), VPS34, VPS15, and ATG14 proteins. Several pieces of evidence indicate that the phosphorylation and ubiquitination of BECN1 at an array of residues fine-tune the responses to diverse autophagy modulating stimuli and helps in maintaining the balance between pro-survival autophagy and pro-apoptotic responses. In this mini-review, we will discuss the importance of distinct BECN1 phosphorylation events, the diverse signaling pathways and kinases involved and their role in the regulation of autophagy.

TNF expression of macrophages is under stringent translational control that depends on the p38 MAPK/MK2 pathway and the AU–rich element (ARE) in the TNF mRNA. Here, we elucidate the molecular mechanism of phosphorylation-regulated translation of TNF. We demonstrate that translation of the TNF-precursor at the ER requires expression of the ARE–binding and -stabilizing factor human antigen R (HuR) together with either activity of the p38 MAPK/MK2 pathway or the absence of the ARE-binding and -destabilizing factor tristetraprolin (TTP). We show that phosphorylation of TTP by MK2 decreases its affinity to the ARE, inhibits its ability to replace HuR, and permits HuR-mediated initiation of translation of TNF mRNA. Since translation of TTP's own mRNA is also regulated by this mechanism, an intrinsic feedback control of the inflammatory response is ensured. The phosphorylation-regulated TTP/HuR exchange at target mRNAs provides a reversible switch between unstable/non-translatable and stable/efficiently translated mRNAs. For immediate response and better control of gene expression, eukaryotic cells have developed means to specifically regulate the stability and translation of pre-formed mRNA transcripts. This post-transcriptional regulation of gene expression is realized by a variety of mRNA-binding proteins, which target specific mRNA sequence elements in a signal-dependent manner. Here we describe a molecular switch mechanism where the exchange of two mRNA-binding proteins is regulated by stress and inflammatory signals. This switch operates between stabilization and efficient translation of the target mRNA, when the activator protein of translational initiation binds instead of the phosphorylated destabilizing protein, and translational arrest and degradation of the target, when the non-phosphorylated destabilizing protein replaces the activator. This mechanism is specific to the mRNA of the inflammatory cytokine tumor necrosis factor (TNF)-α and the mRNA of its regulator protein TTP and, hence, enables fast inflammatory response and its stringent feedback control.

MRNA stability is a major determinant of inflammatory gene expression. Rapid degradation of interleukin-8 (IL-8) mRNA is imposed by a bipartite AU-rich element (ARE) in the 3' untranslated region (R. Winzen et al., Mol. Cell. Biol. 24:4835-4847, 2004). Small interfering RNA-mediated knockdown of the ARE-binding protein KSRP resulted in stabilization of IL-8 mRNA or of a beta-globin reporter mRNA containing the IL-8 ARE. Rapid deadenylation was impaired, indicating a crucial role for KSRP in this step of mRNA degradation. The two IL-8 ARE domains both contribute to interaction with KSRP, corresponding to the importance of both domains for rapid degradation. Exposure to the inflammatory cytokine IL-1 has been shown to stabilize IL-8 mRNA through p38 mitogen-activated protein (MAP) kinase and MK2. IL-1 treatment impaired the interaction of KSRP with the IL-8 ARE in a manner dependent on p38 MAP kinase but apparently independent of MK2. Instead, evidence that TTP, a target of MK2, can also destabilize the IL-8 ARE reporter mRNA is presented. In a comprehensive approach to identify mRNAs controlled by KSRP, two criteria were evaluated by microarray analysis of (i) association of mRNAs with KSRP in pulldown assays and (ii) increased amounts in KSRP knockdown cells. According to both criteria, a group of 100 mRNAs is controlled by KSRP, many of which are unstable and encode proteins involved in inflammation. These results indicate that KSRP functions as a limiting factor in inflammatory gene expression.

The total number of protein-coding genes in the human genome is not significantly higher than those in much simpler eukaryotes, despite a general increase in genome size proportionate to the organismal complexity. The large non-coding transcriptome and extensive differential splicing, are increasingly being accepted as the factors contributing to the complex mammalian physiology and architecture. Recent studies reveal additional layers of functional complexity: some long non-coding RNAs have been re-defined as micropeptide or microprotein encoding transcripts, and in turn some protein-coding RNAs are bifunctional and display also non-coding functions. Moreover, several protein-coding genes express long non-coding RNA splice-forms and generate circular RNAs in addition to their canonical mRNA transcripts, revoking the strict definition of a gene as coding or non-coding. In this mini review, we discuss the current understanding of these hybrid genes and their possible roles and relevance.

Post-transcriptional mechanisms play an important role in the control of inflammatory gene expression. The heterogeneous nuclear ribonucleoprotein K homology (KH)-type splicing regulatory protein (KSRP) triggers rapid degradation of mRNAs for various cytokines, chemokines, and other inflammation-related proteins by interacting with AU-rich elements (AREs) in the 3'-untranslated mRNA regions. In addition to destabilizing mRNAs, AU-rich elements can restrict their translation. Evidence that KSRP also participates in translational silencing was obtained in a screen comparing the polysome profiles of cells with siRNA-mediated depletion of KSRP with that of control cells. Among the group of mRNAs showing increased polysome association upon KSRP depletion are those of interleukin (IL)-6 and IL-1α as well as other ARE-containing transcripts. Redistribution of IL-6 mRNA to polysomes was associated with increased IL-6 protein secretion by the KSRP-depleted cells. Silencing of IL-6 and IL-1α mRNAs depended on their 3'-untranslated regions. The sequence essential for translational control of IL-6 mRNA and its interaction with KSRP was located to an ARE. KSRP-dependent silencing was reversed by IL-1, a strong inducer of IL-6 mRNA and protein expression. The results identify KSRP as a protein involved in ARE-mediated translational silencing. They suggest that KSRP restricts inflammatory gene expression not only by enhancing degradation of mRNAs but also by inhibiting translation, both functions that are counteracted by the proinflammatory cytokine IL-1.