Haploinsufficiency of the 22q11.2 microdeletion gene Mrpl40 disrupts short-term synaptic plasticity and working memory through dysregulation of mitochondrial calcium.

Hemizygous deletion of a 1.5- to 3-megabase region on chromosome 22 causes 22q11.2 deletion syndrome (22q11DS), which constitutes one of the strongest genetic risks for schizophrenia. Mouse models of 22q11DS have abnormal short-term synaptic plasticity that contributes to working-memory deficiencies similar to those in schizophrenia. We screened mutant mice carrying hemizygous deletions of 22q11DS genes and identified haploinsufficiency of Mrpl40 (mitochondrial large ribosomal subunit protein 40) as a contributor to abnormal short-term potentiation (STP), a major form of short-term synaptic plasticity. Two-photon imaging of the genetically encoded fluorescent calcium indicator GCaMP6, expressed in presynaptic cytosol or mitochondria, showed that Mrpl40 haploinsufficiency deregulates STP via impaired calcium extrusion from the mitochondrial matrix through the mitochondrial permeability transition pore. This led to abnormally high cytosolic calcium transients in presynaptic terminals and deficient working memory but did not affect long-term spatial memory. Thus, we propose that mitochondrial calcium deregulation is a novel pathogenic mechanism of cognitive deficiencies in schizophrenia.

Characterization of RIPK3-mediated phosphorylation of the activation loop of MLKL during necroptosis.

Mixed lineage kinase domain-like pseudokinase (MLKL) mediates necroptosis by translocating to the plasma membrane and inducing its rupture. The activation of MLKL occurs in a multimolecular complex (the 'necrosome'), which is comprised of MLKL, receptor-interacting serine/threonine kinase (RIPK)-3 (RIPK3) and, in some cases, RIPK1. Within this complex, RIPK3 phosphorylates the activation loop of MLKL, promoting conformational changes and allowing the formation of MLKL oligomers, which migrate to the plasma membrane. Previous studies suggested that RIPK3 could phosphorylate the murine MLKL activation loop at Ser345, Ser347 and Thr349. Moreover, substitution of the Ser345 for an aspartic acid creates a constitutively active MLKL, independent of RIPK3 function. Here we examine the role of each of these residues and found that the phosphorylation of Ser345 is critical for RIPK3-mediated necroptosis, Ser347 has a minor accessory role and Thr349 seems to be irrelevant. We generated a specific monoclonal antibody to detect phospho-Ser345 in murine cells. Using this antibody, a series of MLKL mutants and a novel RIPK3 inhibitor, we demonstrate that the phosphorylation of Ser345 is not required for the interaction between RIPK3 and MLKL in the necrosome, but is essential for MLKL translocation, accumulation in the plasma membrane, and consequent necroptosis.

HCV infection induces mitochondrial bioenergetic unbalance: causes and effects.

Cells infected by the hepatitis C virus (HCV) are characterized by endoplasmic reticulum stress, deregulation of the calcium homeostasis and unbalance of the oxido-reduction state. In this context, mitochondrial dysfunction proved to be involved and is thought to contribute to the outcome of the HCV-related disease. Here, we propose a temporal sequence of events in the HCV-infected cell whereby the primary alteration consists of a release of Ca(2+) from the endoplasmic reticulum, followed by uptake into mitochondria. This causes successive mitochondrial alterations comprising generation of reactive oxygen and nitrogen species and impairment of the oxidative phosphorylation. A progressive adaptive response results in an enhancement of the glycolytic metabolism sustained by up-regulation of the hypoxia inducible factor. Pathogenetic implications of the model are discussed.

Coexistence of mutations in PINK1 and mitochondrial DNA in early onset parkinsonism.

AIMS AND BACKGROUND: Various genes have been identified for monogenic disorders resembling Parkinson's disease. The products of some of these genes are associated with mitochondria and have been implicated in cellular protection against oxidative damage. In the present study we analysed fibroblasts from a patient carrying the homozygous mutation p.W437X in the PTEN-induced kinase 1 (PINK1), which manifested a very early onset parkinsonism. RESULTS: Patient's fibroblasts did not show variation in the mtDNA copy number or in the expression of the oxidative phosphorylation complexes. Sequence analysis of the patient's mtDNA presented two new missense mutations in the ND5 (m.12397A>G, p.T21A) and ND6 (m. 14319T>C, p.N119D) genes coding for two subunits of complex I. The two mutations were homoplasmic in both the patient and the patient's mother. Patient's fibroblasts resulted in enhanced constitutive production of the superoxide anion radical that was abrogated by inhibitor of the complex I. Moreover enzyme kinetic analysis of the NADH:ubiquinone oxidoreductase showed changes in the substrates affinity. CONCLUSION: To our knowledge, this is the first report showing co-segregation of a Parkinson's disease related nuclear gene mutation with mtDNA mutation(s). Our observation might shed light on the clinical heterogeneity of the hereditary cases of Parkinson's disease, highlighting the hitherto unappreciated impact of coexisting mtDNA mutations in determining the development and the clinical course of the disease.