f Mitochondria possess an outer membrane (OMM) and an inner membrane (IMM), which folds into invaginations called cristae. Lipid composition, membrane potential, and proteins in the IMM influence organization of cristae. Here we show an essential role of the OMM protein Sam50 in the maintenance of the structure of cristae. Sam50 is a part of the sorting and assembly machinery (SAM) necessary for the assembly of ␤-barrel proteins in the OMM. We provide evidence that the SAM components exist in a large protein complex together with the IMM proteins mitofilin and CHCHD3, which we term the mitochondrial intermembrane space bridging (MIB) complex. Interactions between OMM and IMM components of the MIB complex are crucial for the preservation of cristae. After destabilization of the MIB complex, we observed deficiency in the assembly of respiratory chain complexes. Long-term depletion of Sam50 influences the amounts of proteins from all large respiratory complexes that contain mitochondrially encoded subunits, pointing to a connection between the structural integrity of cristae, assembly of respiratory complexes, and/or the maintenance of mitochondrial DNA (mtDNA).

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BackgroundThe ArsRS two-component system is the master regulator of acid adaptation in the human gastric pathogen Helicobacter pylori. Low pH is supposed to trigger the autophosphorylation of the histidine kinase ArsS and the subsequent transfer of the phosphoryl group to its cognate response regulator ArsR which then acts as an activator or repressor of pH-responsive genes. Orthologs of the ArsRS two-component system are also present in H. pylori's close relatives H. hepaticus, Campylobacter jejuni and Wolinella succinogenes which are non-gastric colonizers.Methodology/Principal FindingsIn order to investigate the mechanism of acid perception by ArsS, derivatives of H. pylori 26695 expressing ArsS proteins with substitutions of the histidine residues present in its periplasmic input domain were constructed. Analysis of pH-responsive transcription of selected ArsRS target genes in these mutants revealed that H94 is relevant for pH sensing, however, our data indicate that protonatable amino acids other than histidine contribute substantially to acid perception by ArsS. By the construction and analysis of H. pylori mutants carrying arsS allels from the related ε-proteobacteria we demonstrate that WS1818 of W. succinogenes efficiently responds to acidic pH.Conclusions/SignificanceWe show that H94 in the input domain of ArsS is crucial for acid perception in H. pylori 26695. In addition our data suggest that ArsS is able to adopt different conformations depending on the degree of protonation of acidic amino acids in the input domain. This might result in different activation states of the histidine kinase allowing a gradual transcriptional response to low pH conditions. Although retaining considerable similarity to ArsS the orthologous proteins of H. hepaticus and C. jejuni may have evolved to sensors of a different environmental stimulus in accordance with the non gastric habitat of these bacteria.

As a consequence of their bacterial origin, mitochondria contain ␤-barrel proteins in their outer membrane (OMM). These proteins require the translocase of the outer membrane (TOM) complex and the conserved sorting and assembly machinery (SAM) complex for transport and integration into the OMM. The SAM complex and the ␤-barrel assembly machinery (BAM) required for biogenesis of ␤-barrel proteins in bacteria are evolutionarily related. Despite this homology, we show that bacterial ␤-barrel proteins are not universally recognized and integrated into the OMM of human mitochondria. Selectivity exists both at the level of the TOM and the SAM complex. Of all of the proteins we tested, human mitochondria imported only ␤-barrel proteins originating from Neisseria sp., and only Omp85, the central component of the neisserial BAM complex, integrated into the OMM. PorB proteins from different Neisseria, although imported by the TOM, were not recognized by the SAM complex and formed membrane complexes only when functional Omp85 was present at the same time in mitochondria. Omp85 alone was capable of integrating other bacterial ␤-barrel proteins in human mitochondria, but could not substitute for the function of its mitochondrial homolog Sam50. Thus, signals and machineries for transport and assembly of ␤-barrel proteins in bacteria and human mitochondria differ enough to allow only a certain type of ␤-barrel proteins to be targeted and integrated in mitochondrial membranes in human cells.Mitochondria are organelles of bacterial origin, surrounded by an outer (OMM) 4 and an inner membrane (IMM). The OMM contains ␤-barrel proteins, a class of pore-forming proteins additionally found only in chloroplasts and Gram-negative bacteria (1, 2). Similar to the majority of other mitochondrial proteins, ␤-barrel proteins are synthesized in the cytosol and have to be imported into mitochondria with the help of the translocase of the outer mitochondrial membrane (TOM) complex. For the membrane integration and assembly into complexes, ␤-barrel proteins require additional proteinaceous machinery in the OMM. This is so called sorting and assembly machinery (SAM), also known as topogenesis of outer membrane ␤-barrel proteins (TOB complex) (3, 4).The central component of the SAM complex is Sam50/ Tob55, a protein with a function that has been conserved from bacteria to human (4 -6). Other components of the SAM complex include the Sam35/Tob38/Tom38 (7-9) and Sam37/ Mas37/Tom37 (3, 10) proteins in yeast, and Metaxin 1 and Metaxin 2 (11) in mammalian mitochondria.The signals in ␤-barrel proteins that are recognized by the TOM complex belong to internal targeting signals and are not yet fully understood. However, a specific signal has been identified in the C-terminal part of mitochondrial ␤-barrel proteins that directs them to the SAM complex to be properly sorted and integrated into the OMM (12). Unlike in yeast, this signal in mammalian ␤-barrel proteins is always present at the extreme C terminus of the protein, and the addition of even a shor...

Awareness of estrogen's neuroprotective and behavioral effects is broadening rapidly and has served as an incentive to investigate estrogen signaling in central nervous system disorders. The present analysis focuses on two human nuclear estrogen receptors, ER alpha and ER beta, which have been shown to play key roles in the complex integration of estrogen's genomic and non-genomic modes of action. The corresponding genes are estimated to have diverged from an ancestral ER gene over 450 million years ago and are candidate genes for a variety of brain disorders. Recent progress in the Human Genome Project has greatly aided our understanding of the molecular blueprint and provides the means for reassessing both genes' genomic organization. Analyses of multiple alternatively spliced transcripts, large untranslated sequences and neighbouring genes reveal several novel features which suggest an increasingly versatile transcriptional machinery. We outline additional exons in the genes' 5'- and 3'-untranslated regions, a new polymorphic ER alpha microsatellite and a nested gene which lend themselves to further evolutionary and functional studies.