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Parvalbumin‐positive (PV+) fast‐spiking interneurons are essential to control the firing activity of principal neuron ensembles, thereby regulating cognitive processes. The high firing frequency activity of PV+ interneurons imposes high‐energy demands on their metabolism that must be supplied by distinctive machinery for energy generation. Exploring single‐cell transcriptomic data for the mouse cortex, we identified a metabolism‐associated gene with highly restricted expression to PV+ interneurons: Cox6a2, which codes for an isoform of a cytochrome c oxidase subunit. Cox6a2 deletion in mice disrupts perineuronal nets and enhances oxidative stress in PV+ interneurons, which in turn impairs the maturation of their morphological and functional properties. Such dramatic effects were likely due to an essential role of COX6A2 in energy balance of PV+ interneurons, underscored by a decrease in the ATP‐to‐ADP ratio in Cox6a2−/−PV+ interneurons. Energy disbalance and aberrant maturation likely hinder the integration of PV+ interneurons into cortical neuronal circuits, leading to behavioral alterations in mice. Additionally, in a human patient bearing mutations in COX6A2, we found a potential association of the mutations with mental/neurological abnormalities. Synopsis: Whether brain neurons firing at high frequency depend on distinct metabolic machineries to satisfy their increased energy demands remains unclear. Here, combined genetic, functional and behavioural analyses reveal a selective role for the oxidative phosphorylation in stress protection of interneuron by safeguarding their energy supply. Mitochondrial complex IV subunit isoform Cox6a2 is enriched in parvalbumin‐positive (PV+) interneurons in the adult rodent, rhesus monkey, and human brain.PV+ interneuron‐specific Cox6a2 ablation decreases cellular ATP levels and increases oxidative stress in mice.Cox6a2 loss impairs transcriptional and morphological maturation during postnatal stages.Adult Cox6a2‐depleted PV+ interneurons are hyperexcitable and defective in repetitive firing.Cox6a2 depletion results in hyperactivity in vivo. [ABSTRACT FROM AUTHOR]

Abstract: Rationale: Experienced primary care physicians handle most illnesses to everyone's satisfaction despite limited resources of time and means. However, cases can be multifaceted in that harmless‐presenting symptoms may also be warning signals or an indicator of a health disorder that too infrequently presents in family practice to be diagnosed correctly. On the basis of these observations, RN Braun developed 82 diagnostic protocols for a structured recording of various complaints. Method: All consultations during the years 2001 to 2014, in which 1 author (WF) had used diagnostic protocols in her single‐handed practice, were analyzed retrospectively regarding reasons for encounter, diagnostic classification, and long‐term outcome. Results: During the period, a diagnostic protocol was used 1686 times. It was applied at a rate of approximately 5% of 2500 new complaints annually, most often (1366 times) for febrile conditions. In 320 consultations for other complaints, 43 different diagnostic protocols were applied. Among them, the “tabula diagnostica” for various undifferentiated symptoms was used most frequently (n = 54), followed by diagnostic protocols for headache (n = 45), dizziness (n = 36), precordial pain (n = 20), nonspecific abdominal pain (n = 15), low back pain (n = 14), hypertension (n = 12), diarrhea > 1 week (n = 12), epigastralgia (n = 11), depression (n = 10), polyarthralgia (n = 8), cough, and lower abdominal pain (each n = 7). A final diagnosis was established in less than 20% of cases. Conclusions: This observational study from routine practice gives an insight how diagnostic protocols helped to manage complex patient presentations. A broader use of diagnostic protocols could investigate the potential of this consultation tool to handle the complexity of primary health care. The use of a standardized diagnostic approach could stimulate research, in particular on managing common complaints/undifferentiated illness and their inherent diagnostic uncertainty. [ABSTRACT FROM AUTHOR]

The RNA-directed DNA methylation (Rd DM) pathway is of central importance to the initiation and maintenance of transcriptional gene silencing in plants. DNA methylation is directed to target sequences by a mechanism that involves production of small RNAs by RNA polymerase IV and long non-coding RNAs by RNA polymerase V. DNA methylation then leads to recruitment of histone-modifying enzymes, followed by establishment of a silenced chromatin state. Recently MORC6, a member of the microrchidia ( MORC) family of adenosine triphosphatases ( ATPases), has been shown to be involved in transcriptional gene silencing. However, reports differ regarding whether MORC6 is involved in Rd DM itself or acts downstream of DNA methylation to enable formation of higher-order chromatin structure. Here we demonstrate that MORC6 is required for efficient Rd DM at some target loci, and, using a GFP reporter system, we found that morc6 mutants show a stochastic silencing phenotype. By using cell sorting to separate silenced and unsilenced cells, we show that release of silencing at this locus is associated with a loss of DNA methylation. Thus our data support a view that MORC6 influences Rd DM and that it is not acting downstream of DNA methylation. For some loci, efficient initiation or maintenance of DNA methylation may depend on the ability to form higher-order chromatin structure. [ABSTRACT FROM AUTHOR]

Higher-order multi-protein complexes such as RNA polymerase II (Pol II) complexes with transcription initiation factors are often not amenable to X-ray structure determination. Here, we show that protein cross-linking coupled to mass spectrometry (MS) has now sufficiently advanced as a tool to extend the Pol II structure to a 15-subunit, 670 kDa complex of Pol II with the initiation factor TFIIF at peptide resolution. The N-terminal regions of TFIIF subunits Tfg1 and Tfg2 form a dimerization domain that binds the Pol II lobe on the Rpb2 side of the active centre cleft near downstream DNA. The C-terminal winged helix (WH) domains of Tfg1 and Tfg2 are mobile, but the Tfg2 WH domain can reside at the Pol II protrusion near the predicted path of upstream DNA in the initiation complex. The linkers between the dimerization domain and the WH domains in Tfg1 and Tfg2 are located to the jaws and protrusion, respectively. The results suggest how TFIIF suppresses non-specific DNA binding and how it helps to recruit promoter DNA and to set the transcription start site. This work establishes cross-linking/MS as an integrated structure analysis tool for large multi-protein complexes. [ABSTRACT FROM AUTHOR]