In mammals, most physiology and behavior are subject to well-controlled daily oscillations. Thus, sleep-wake cycles, heartbeat frequency, blood pressure, body temperature, renal activity, liver metabolism, and the secretion of many hormones are controlled by an endogenous time measuring system called the circadian clock (Portaluppi et

Communication between organelles is an important feature of all eukaryotic cells. To uncover components involved in mitochondria/endoplasmic reticulum (ER) junctions, we screened for mutants that could be complemented by a synthetic protein designed to artificially tether the two organelles. We identified the Mmm1/Mdm10/Mdm12/Mdm34 complex as a molecular tether between ER and mitochondria. The tethering complex was composed of proteins resident of both ER and mitochondria. With the use of genome-wide mapping of genetic interactions, we showed that the components of the tethering complex were functionally connected to phospholipid biosynthesis and calcium-signaling genes. In mutant cells, phospholipid biosynthesis was impaired. The tethering complex localized to discrete foci, suggesting that discrete sites of close apposition between ER and mitochondria facilitate interorganelle calcium and phospholipid exchange.Eukaryotic cells evolved segregation of functions into separate organelles. Compartmentalization increases the efficiency of biochemical reactions by creating tailored chemical microenvironments, but also creates a need for communication and routes of metabolite exchange. Membrane lipids, for example, are primarily synthesized in the endoplasmic reticulum (ER) and distributed to other organelles. Many organelles exchange phospholipids with the ER via vesicular transport. In contrast, mitochondria are not connected to vesicular trafficking pathways, and many lipids of the inner and outer mitochondrial Other work has implicated ER-mitochondrial contact sites in Ca ++ transport between the ER and mitochondria (4-6), suggesting a mechanism that may exploit the formation of an encapsulated space at the contact sites, akin to that formed at neuronal or immunological synapses. Such a connection between the ER and the mitochondria might buffer and control cytosolic and mitochondrial Ca ++ concentrations (7). Several proteins have been implicated to participate in ER-mitochondria contacts, including the ER resident Ca ++ channel IP3 receptor, the mitochondrial voltage-dependent anion channel, the chaperones grp75 and sigma-1R, the sorting protein PACS-2, and the mitofusin Mfn2 (8-11).To explore a role for ER-mitochondrial junctions, we sought mutants in the yeast Saccharomyces cerevisiae, in which tethering between the two organelles was impaired. We reasoned that, if such contacts are important, defects in proteins that establish these interactions would be detrimental, yet perhaps could be suppressed by artificially tethering ER and mitochondria (Fig. 1A). We designed a synthetic ER-mitochondria tether ("ChiMERA" for construct helping in mitochondria-ER association) (Fig. 1B) consisting of an N-terminal mitochondrial signal sequence and transmembrane domain derived from Tom70, a central module composed of green fluorescent protein (GFP), and a C-terminal ER tail-anchor derived from Ubc6. The design was based on a similar chimeric protein that strengthened mitochondria-ER interactions (12). The GFP moiet...

The mammalian circadian timing system consists of a master pacemaker in neurons of the suprachiasmatic nucleus (SCN) and clocks of a similar molecular makeup in most peripheral body cells. Peripheral oscillators are self-sustained and cell autonomous, but they have to be synchronized by the SCN to ensure phase coherence within the organism. In principle, the rhythmic expression of genes in peripheral organs could thus be driven not only by local oscillators, but also by circadian systemic signals. To discriminate between these mechanisms, we engineered a mouse strain with a conditionally active liver clock, in which REV-ERBα represses the transcription of the essential core clock gene Bmal1 in a doxycycline-dependent manner. We examined circadian liver gene expression genome-wide in mice in which hepatocyte oscillators were either running or arrested, and found that the rhythmic transcription of most genes depended on functional hepatocyte clocks. However, we discovered 31 genes, including the core clock gene mPer2, whose expression oscillated robustly irrespective of whether the liver clock was running or not. By contrast, in liver explants cultured in vitro, circadian cycles of mPer2::luciferase bioluminescence could only be observed when hepatocyte oscillators were operational. Hence, the circadian cycles observed in the liver of intact animals without functional hepatocyte oscillators were likely generated by systemic signals. The finding that rhythmic mPer2 expression can be driven by both systemic cues and local oscillators suggests a plausible mechanism for the phase entrainment of subsidiary clocks in peripheral organs.

Close proximities between organelles have been described for decades. However, only recently a specific field dealing with organelle communication at membrane contact sites has gained wide acceptance, attracting scientists from multiple areas of cell biology. The diversity of approaches warrants a unified vocabulary for the field. Such definitions would facilitate laying the foundations of this field, streamlining communication and resolving semantic controversies. This opinion, written by a panel of experts in the field, aims to provide this burgeoning area with guidelines for the experimental definition and analysis of contact sites. It also includes suggestions on how to operationally and tractably measure and analyze them with the hope of ultimately facilitating knowledge production and dissemination within and outside the field of contact-site research.

Membrane-bound organelles are a wonderful evolutionary acquisition of the eukaryotic cell, allowing the segregation of sometimes incompatible biochemical reactions into specific compartments with tailored microenvironments. On the flip side, these isolating membranes that crowd the interior of the cell, constitute a hindrance to the diffusion of metabolites and information to all corners of the cell. To ensure coordination of cellular activities, cells use a network of contact sites between the membranes of different organelles. These membrane contact sites (MCSs) are domains where two membranes come to close proximity, typically less than 30nm. Such contacts create microdomains that favor exchange between two organelles. MCSs are established and maintained in durable or transient states by tethering structures, which keep the two membranes in proximity, but fusion between the membranes does not take place. Since the endoplasmic reticulum (ER) is the most extensive cellular membrane network, it is thus not surprising to find the ER involved in most MCSs within the cell. The ER contacts diverse compartments such as mitochondria, lysosomes, lipid droplets, the Golgi apparatus, endosomes and the plasma membrane. In this review, we will focus on the common organizing principles underlying the many MCSs found between the ER and virtually all compartments of the cell, and on how the ER establishes a network of MCSs for the trafficking of vital metabolites and information. This article is part of a Special Issue entitled: Functional and structural diversity of endoplasmic reticulum.