bioRxiv Science⌕ Search

Biology subjects

Breker, M.

Publications and source records attributed to Breker, M..

7 recordsLinked to original sources

Conserved inhibitory mechanisms of root-associated bacteria towardsArabidopsis thaliana and Chlamydomonas reinhardtii

Soil bacteria colonize plants without causing visible disease, yet many suppress host growth. The prevalence and evolutionary reach of this cryptic inhibition are unknown, because phenotyping hundreds of isolates in plants is prohibitively slow. Here we use the unicellular algae Chlamydomonas reinhardtii as a scalable proxy for the green lineage. We screened 148 plant-associated bacteria on Chlamydomonas lawns. Seven of eight Chlamydomonas inhibitors also inhibited the vascular model plant Arabidopsis thaliana. In one of these dual inhibitors, Burkholderia sola MF6, transposon screening and proteomics implicated type VI secretion and tight-adherence pili, and over half the non-inhibitory mutants also failed against Arabidopsis. In the algae, infection triggers rapid deflagellation, then non-lytic regulated cell death. The algal zinc/iron transporter ZIP3 promotes inhibition, whereas the cathepsin X protease CEP12 restrains it, and ZIP3 is epistatic to CEP12. Cryptic inhibition is thus ancient and widespread, and a unicellular algae uncovers it at scale.

plant biology↗

The ribosome-associated quality control factor Vms1 protects mitochondrial import and homeostasis during translation stress

Mitochondrial protein synthesis and import are tightly coordinated to maintain cellular proteostasis, yet how cytosolic translation stress affects mitochondrial homeostasis remains poorly understood. Here, we investigated the cellular consequences of general translation stress using low-dose translation inhibitors in Saccharomyces cerevisiae. Genome-wide phenotypic screening revealed that the deletion of tRNA-hydrolase VMS1 involved in mitochondria-associated ribosomal quality control (mitoRQC) causes a unique hypersensitivity to low-dose cycloheximide. Quantitative proteomics demonstrated that translation stress triggers specific depletion of mitochondrial proteins, reduced respiratory capacity, and impaired mitochondrial membrane potential. Most notably, cytosolic translation stress strongly inhibited mitochondrial protein import, an effect that was substantially exacerbated in vms1{Delta} cells. We further identify the mitochondrial AAA-ATPase Msp1 as a protective factor during translation stress. Msp1 expression increased in vms1{Delta} cells, and its overexpression restored growth under stress, linking co-translational quality control to mitochondrial protein import surveillance. Together, our findings reveal mitochondrial protein import as a major target of cytosolic translation stress and uncover functional cooperation between mitoRQC and Msp1 in safeguarding mitochondrial proteostasis and cell survival.

cell biology↗

Expanding The Algal Hydrogen Toolbox: A Non-GMO Platform Reveals Multiple Physiological Routes To Sustained Hydrogen Production Across Microalgae

Sustainable hydrogen production from microalgae remains limited by intrinsic physiological constraints and the need to preserve biomass value for food and feed applications. Transgenic approaches to overcome these limitations were proven successful, yet result in genetically modified (GMO) strains that face major regulatory and deployment barriers. Here, we present a non-GMO experimental platform that enables systematic isolation of hydrogen-producing phenotypes through high-throughput UV mutagenesis pipline coupled with targeted physiological screening. Applying this approach across phylogenetically distinct algal species, including the industrial strain Chlorella vulgaris and the extremophile Chlorella ohadii, we achieve high discovery efficiency, recovering 0.4-0.6% validated hydrogen-producing mutants and achieving 6.7-25% validation rates among screen-positive candidates, indicating strong enrichment at the primary screening stage. We show that sustained hydrogen production represents a physiologically accessible state emerging across diverse genetic backgrounds. This state is consistently associated with reorganization of photosynthetic electron partitioning, yet arises through multiple distinct configurations that differentially balance hydrogen production, oxygen metabolism, and carbon fixation. This framework provides a scalable route to identify hydrogen-producing strains in industrially relevant algae without introducing foreign DNA and expands the accessible design space for photobiological hydrogen production.

plant biology↗

Systematic identification and characterization of novel genes in the regulation and biogenesis of photosynthetic machinery

Photosynthesis is central to food production and the Earths biogeochemistry, yet the molecular basis for its regulation remains poorly understood. Here, using high-throughput genetics in the model eukaryotic alga Chlamydomonas reinhardtii, we identify with high confidence (FDR<0.11) 70 previously-uncharacterized genes required for photosynthesis. We then provide a resource of mutant proteomes that enables functional characterization of these novel genes by revealing their relationship to known genes. The data allow assignment of 34 novel genes to the biogenesis or regulation of one or more specific photosynthetic complexes. Additional analysis uncovers at least seven novel critical regulatory proteins, including five Photosystem I mRNA maturation factors and two master regulators: MTF1, which impacts chloroplast gene expression directly; and PMR1, which impacts expression via nuclear-expressed factors. Our work provides a rich resource identifying novel regulatory and functional genes and placing them into pathways, thereby opening the door to a system-level understanding of photosynthesis. HighlightsO_LIHigh-confidence identification of 70 previously-uncharacterized genes required for photosynthesis C_LIO_LIProteomic analysis of mutants allows assignment of function to novel genes C_LIO_LICharacterization of 5 novel Photosystem I mRNA maturation factors validates this resource C_LIO_LIMTF1 and PMR1 identified as master regulators of photosynthesis C_LI

cell biology↗

AMPK regulates ESCRT-dependent microautophagy of proteasomes concomitant with proteasome storage granule assembly during glucose starvation

The ubiquitin-proteasome system regulates numerous cellular processes and is central to protein homeostasis. In proliferating yeast and many mammalian cells, proteasomes are highly enriched in the nucleus. In carbon-starved yeast, proteasomes migrate to the cytoplasm and collect in phase-separated proteasome storage granules (PSGs). PSGs dissolve and proteasomes return to the nucleus within minutes of glucose refeeding. The mechanisms by which cells regulate proteasome homeostasis under these conditions remain largely unknown. Here we show that AMP-activated protein kinase (AMPK) together with endosomal sorting complexes required for transport (ESCRTs) drive a glucose starvation-dependent microautophagy pathway that preferentially sorts aberrant proteasomes into the vacuole, thereby biasing accumulation of functional proteasomes in PSGs. The proteasome core particle (CP) and regulatory particle (RP) are regulated differently. Without AMPK, the insoluble protein deposit (IPOD) serves as an alternative site that specifically sequesters CP aggregates. Our findings reveal a novel AMPK-controlled ESCRT-mediated microautophagy mechanism in the regulation of proteasome trafficking and homeostasis under carbon starvation.

cell biology↗

Large-scale rewiring in a yeast hybrid

The merging of genomes in inter-specific hybrids can result in novel phenotypes, including increased growth rate and biomass yield, a phenomenon known as heterosis. We describe a budding yeast hybrid that grows faster than its parents under different environments. Phenotypically, the hybrid progresses more rapidly through cell cycle checkpoints, relieves the repression of respiration in fast growing conditions, does not slow down its growth when presented with ethanol stress, and shows increasing signs of DNA damage. A systematic genetic screen identified hundreds of alleles affecting hybrid growth whose identity vastly differed between the hybrid and its parent and between growth conditions. This large-scale rewiring of allele effects suggests that despite showing clear heterosis, the hybrid is perturbed in multiple regulatory processes. We discuss the possibility that incompatibilities contribute to hybrid vigor by perturbing safeguard mechanisms that limit growth in the parental background.

systems biology↗

Validated Bayesian differentiation of causative and passenger mutations

In many contexts, the problem arises of determining which of many candidate mutations is the most likely to be causative for some phenotype. It is desirable to have a way to evaluate this probability that relies as little as possible on previous knowledge, to avoid bias against discovering new genes or functions. We are isolating mutants with blocked cell cycle progression in Chlamydomonas, and determining mutant genome sequences. Due to the intensity of UV mutagenesis required for efficient mutant collection, the mutants contain multiple mutations altering coding sequence. To provide a quantitative estimate of probability that each individual mutation in a given mutant is the causative one, we develop a Bayesian approach. The approach employs four independent indicators: sequence conservation of the mutated coding sequence with Arabidopsis; severity of the mutation relative to Chlamydomonas wild type based on Blosum62 scores; meiotic mapping information for location of the causative mutation relative to known molecular markers; and, for a subset of mutants, transcriptional profile of the candidate wild type genes through the mitotic cell cycle.\n\nThese indicators are statistically independent, and so can be combined quantitatively into a single probability calculation. We validate this calculation: recently isolated mutations that were not in the training set for developing the indicators, with high calculated probability of causality, are confirmed in every case by additional genetic data to indeed be causative. Analysis of best reciprocal blast relationships among Chlamydomonas and other eukaryotes indicate that the Ts-lethal mutants that our procedure recovers are highly enriched for fundamental cell-essential functions conserved broadly across plants and other eukaryotes, accounting for the high information content of sequence alignment to Arabidopsis.

genetics↗