Transcriptional silencing through the Polycomb silencing machinery utilizes a "read-write" mechanism involving histone tail modifications. However, nucleation of silencing and long-term stable transmission of the silenced state also requires P-olycomb Repressive Complex 2 (PRC2) accessory proteins, whose molecular role is poorly understood. The Arabidopsis VEL proteins are accessory proteins that interact with PRC2 to nucleate and propagate silencing at the FLOWERING LOCUS C (FLC) locus, enabling early flowering in spring. Here, we report that VEL proteins contain a domain related to an atypical four-helix bundle that engages in spontaneous concentration-dependent head-to-tail polymerization to assemble dynamic biomolecular condensates. Mutations blocking polymerization of this VEL domain prevent Polycomb silencing at FLC. Plant VEL proteins thus facilitate assembly of dynamic multivalent Polycomb complexes required for inheritance of the silenced state.

BACKGROUND:Mitochondria provide ATP through the process of oxidative phosphorylation, physically located in the inner mitochondrial membrane (IMM). The mitochondrial contact site and organising system (MICOS) complex is known as the 'mitoskeleton' due to its role in maintaining IMM architecture. APOO encodes MIC26, a component of MICOS, whose exact function in its maintenance or assembly has still not been completely elucidated.METHODS:We have studied a family in which the most affected subject presented progressive developmental delay, lactic acidosis, muscle weakness, hypotonia, weight loss, gastrointestinal and body temperature dysautonomia, repetitive infections, cognitive impairment and autistic behaviour. Other family members showed variable phenotype presentation. Whole exome sequencing was used to screen for pathological variants. Patient-derived skin fibroblasts were used to confirm the pathogenicity of the variant found in APOO. Knockout models in Drosophila melanogaster and Saccharomyces cerevisiae were employed to validate MIC26 involvement in MICOS assembly and mitochondrial function.RESULTS:A likely pathogenic c.350T>C transition was found in APOO predicting an I117T substitution in MIC26. The mutation caused impaired processing of the protein during import and faulty insertion into the IMM. This was associated with altered MICOS assembly and cristae junction disruption. The corresponding mutation in MIC26 or complete loss was associated with mitochondrial structural and functional deficiencies in yeast and D. melanogaster models.CONCLUSION:This is the first case of pathogenic mutation in APOO, causing altered MICOS assembly and neuromuscular impairment. MIC26 is involved in the assembly or stability of MICOS in humans, yeast and flies.

Moderate overexpression of Opa1, encoding a master regulator of mitochondrial cristae morphology, has been shown to improve significantly mitochondrial damage induced by drugs, surgical denervation, or genetically determined OXPHOS defects. However, this approach has been so far demonstrated in a limited number of genetically defective OXPHOS models characterized by specific impairment of a single mitochondrial respiratory chain complex. Here, we investigated the effectiveness of moderate Opa1 overexpression in the Mpv17-/- mouse, characterized by profound, multisystem mtDNA depletion. In naïve Mpv17-/- individuals, whose genetic background was crossed with individuals belonging to the Opa1tg strain, we found a surprising anticipation of severe, progressive focal segmental glomerulosclerosis, previously described in Mpv17-/- animals as a late-onset clinical feature (after 12-18 months of life). In contrast, kidney failure led Mpv17-/- individuals from this new “mixed” strain leading to death 8-9 weeks after birth. However, Mpv17-/-::Opa1tg mice lived much longer than Mpv17-/- littermates, and developed much later severe proteinuria associated with focal segmental glomerulosclerosis. MtDNA content and OXPHOS activities were significantly higher in Mpv17-/-::Opa1tg than in Mpv17-/- kidneys, and similar to WT littermates. Mitochondrial network and cristae ultrastructure were largely preserved in Mpv17-/-::Opa1tg vs. Mpv17-/- kidney and isolated podocytes. Mechanistically, the protective effect of Opa1 overexpression in this model was mediated by a block in apoptosis due to the stabilization of the mitochondrial cristae, consequently increasing the levels of mitochondrial morphology proteins like MFN2 and MIC19 as well as stabilizing ATP synthase oligomers. These results demonstrate that strategies aiming at increasing Opa1 expression or activity can be an effective aid against mtDNA depletion syndromes.

AbstractModerate overexpression ofOpa1, encoding a master regulator of mitochondrial cristae morphology, has been shown to improve significantly mitochondrial damage induced by drugs, surgical denervation, or genetically determined OXPHOS defects. However, this approach has been so far demonstrated in a limited number of genetically defective OXPHOS models characterized by specific impairment of a single mitochondrial respiratory chain complex. Here, we investigated the effectiveness of moderateOpa1overexpression in theMpv17-/-mouse, characterized by profound, multisystem mtDNA depletion. In naïveMpv17-/-individuals, whose genetic background was crossed with individuals belonging to theOpa1tgstrain, we found a surprising anticipation of severe, progressive focal segmental glomerulosclerosis, previously described inMpv17-/-animals as a late-onset clinical feature (after 12-18 months of life). In contrast, kidney failure ledMpv17-/-individuals from this new “mixed” strain leading to death 8-9 weeks after birth. However,Mpv17-/-::Opa1tgmice lived much longer thanMpv17-/-littermates, and developed much later severe proteinuria associated with focal segmental glomerulosclerosis. MtDNA content and OXPHOS activities were significantly higher inMpv17-/-::Opa1tgthan inMpv17-/-kidneys, and similar to WT littermates. Mitochondrial network and cristae ultrastructure were largely preserved inMpv17-/-::Opa1tgvs.Mpv17-/-kidney and isolated podocytes. Mechanistically, the protective effect ofOpa1overexpression in this model was mediated by a block in apoptosis due to the stabilization of the mitochondrial cristae, consequently increasing the levels of mitochondrial morphology proteins like MFN2 and MIC19 as well as stabilizing ATP synthase oligomers. These results demonstrate that strategies aiming at increasingOpa1expression or activity can be an effective aid against mtDNA depletion syndromes.

The absence of state transitions in a Nt(Hn) cybrid is due to a cleavage of the threonine residue from the misprocessed N-terminus of the LHCII polypeptides. The cooperation between the nucleus and chloroplast genomes is essential for plant photosynthetic fitness. The rapid and specific interactions between nucleus-encoded and chloroplast-encoded proteins are under intense investigation with potential for applications in agriculture and renewable energy technology. Here, we present a novel model for photosynthesis research in which alien henbane (Hyoscyamus niger) chloroplasts function on the nuclear background of a tobacco (Nicotiana tabacum). The result of this coupling is a cytoplasmic hybrid (cybrid) with inhibited state transitions—a mechanism responsible for balancing energy absorption between photosystems. Protein analysis showed differences in the LHCII composition of the cybrid plants. SDS-PAGE analysis revealed a novel banding pattern in the cybrids with at least one additional ‘LHCII’ band compared to the wild-type parental species. Proteomic work suggested that the N-terminus of at least some of the cybrid Lhcb proteins was missing. These findings provide a mechanistic explanation for the lack of state transitions—the N-terminal truncation of the Lhcb proteins in the cybrid included the threonine residue that is phosphorylated/dephosphorylated in order to trigger state transitions and therefore crucial energy balancing mechanism in plants.

Photoprotective non-photochemical quenching (NPQ) represents an effective way to dissipate the light energy absorbed in excess by most phototrophs. It is often claimed that NPQ formation/relaxation kinetics are determined by xanthophyll composition. We, however, found that, for the alveolate alga Chromera velia, this is not the case. In the present paper, we investigated the reasons for the constitutive high rate of quenching displayed by the alga by comparing its light harvesting strategies with those of a model phototroph, the land plant Spinacia oleracea. Experimental results and in silico studies support the idea that fast quenching is due not to xanthophylls, but to intrinsic properties of the Chromera light harvesting complex (CLH) protein, related to amino acid composition and protein folding. The pKa for CLH quenching was shifted by 0.5 units to a higher pH compared with higher plant antennas (light harvesting complex II; LHCII). We conclude that, whilst higher plant LHCIIs are better suited for light harvesting, CLHs are 'natural quenchers' ready to switch into a dissipative state. We propose that organisms with antenna proteins intrinsically more sensitive to protons, such as C. velia, carry a relatively high concentration of violaxanthin to improve their light harvesting. In contrast, higher plants need less violaxanthin per chlorophyll because LHCII proteins are more efficient light harvesters and instead require co-factors such as zeaxanthin and PsbS to accelerate and enhance quenching.

This study, based within the catchment area of the River Frome, an important chalk stream in the south of England, compared ciliated protozoan communities associated with three species of aquatic macrophyte common to lotic habitats: Ranunculus penicillatus subsp. pseudofluitans, Nasturtium officinale and Sparganium emersum. A total of 77 ciliate species were counted. No species-specific ciliate assemblage was found to be typical of any one plant species. Ciliate abundance between plant species was determined to be significantly different. The ciliate communities from each plant species were unique in that the number of species increased with ciliate abundance. The community associated with R. penicillatus subsp. pseudofluitans showed the highest consistency and species richness whereas S. emersum ciliate communities were unstable. Most notably, N. officinale was associated with low ciliate abundances and an apparent reduction in biofilm formation, discussed herein in relation to the plant's production of the microbial toxin phenethyl isothiocyanate. We propose that the results reflect differences in the quantity and quality of biofilm present on the plants, which could be determined by the different plant morphologies, patterns of plant decay and herbivore defense systems, all of which suppress or promote the various conditions for biofilm growth.