RNAIII is the intracellular effector of the quorum-sensing system in Staphylococcus aureus. It is one of the largest regulatory RNAs (514 nucleotides long) that are known to control the expression of a large number of virulence genes. Here, we show that the 3 domain of RNAIII coordinately represses at the post-transcriptional level, the expression of mRNAs that encode a class of virulence factors that act early in the infection process. We demonstrate that the 3 domain acts primarily as an antisense RNA and rapidly anneals to these mRNAs, forming long RNA duplexes. The interaction between RNAIII and the mRNAs results in repression of translation initiation and triggers endoribonuclease III hydrolysis. These processes are followed by rapid depletion of the mRNA pool. In addition, we show that RNAIII and its 3 domain mediate translational repression of rot mRNA through a limited number of base pairings involving two loop-loop interactions. Since Rot is a transcriptional regulatory protein, we proposed that RNAIII indirectly acts on many downstream genes, resulting in the activation of the synthesis of several exoproteins. These data emphasize the multitude of regulatory steps affected by RNAIII and its 3 domain in establishing a network of S. aureus virulence factors.[Keywords: Regulatory RNA; translational repression; antisense regulation; RNase III; virulence; Staphylococcus aureus]Supplemental material is available at http://www.genesdev.org.

Mitochondria-associated endoplasmic reticulum (ER) membranes (MAMs) Mitochondria and endoplasmic reticulum (ER) networks are interconnected, sharing structural and functional interactions essential for the maintenance of cellular homeostasis. The contacts between ER and mitochondria, known as mitochondria-associated ER membranes (MAMs), play a pivotal role in calcium (Ca 2+ ) signaling, lipid transport, energy metabolism, and cell survival (1). The physical interactions between both organelles depend on complementary membrane proteins, which tether the two organelles together at specific sites. For example, the voltage-dependent anion channel (VDAC) of the outer mitochondrial membrane interacts with the inositol 1,4,5-triphosphate receptor (IP3R) on the ER through the molecular chaperone glucoseregulated protein 75 (Grp75), allowing Ca 2+ transfer from the ER to mitochondria (2). Recently, mitofusin 2 (Mfn2)

Modifications of the interactions between endoplasmic reticulum (ER) and mitochondria, defined as mitochondria-associated membranes (MAMs), were recently shown to be involved in the control of hepatic insulin action and glucose homeostasis, but with conflicting results. Whereas skeletal muscle is the primary site of insulin-mediated glucose uptake and the main target for alterations in insulin-resistant states, the relevance of MAM integrity in muscle insulin resistance is unknown. Deciphering the importance of MAMs on muscle insulin signaling could help to clarify this controversy. Here, we show in skeletal muscle of different mice models of obesity and type 2 diabetes (T2D) a marked disruption of ER-mitochondria interactions as an early event preceding mitochondrial dysfunction and insulin resistance. Furthermore, in human myotubes, palmitate-induced insulin resistance is associated with a reduction of structural and functional ER-mitochondria interactions. Importantly, experimental increase of ER-mitochondria contacts in human myotubes prevents palmitate-induced alterations of insulin signaling and action, whereas disruption of MAM integrity alters the action of the hormone. Lastly, we found an association between altered insulin signaling and ER-mitochondria interactions in human myotubes from obese subjects with or without T2D compared with healthy lean subjects. Collectively, our data reveal a new role of MAM integrity in insulin action of skeletal muscle and highlight MAM disruption as an essential subcellular alteration associated with muscle insulin resistance in mice and humans. Therefore, reduced ER-mitochondria coupling could be a common alteration of several insulin-sensitive tissues playing a key role in altered glucose homeostasis in the context of obesity and T2D.