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Clemente, T.

Publications and source records attributed to Clemente, T..

3 recordsLinked to original sources

The multifunction Coxiella effector Vice stimulates macropinocytosis and interferes with the ESCRT machinery

Intracellular bacterial pathogens divert multiple cellular pathways to establish their niche and persist inside their host. Coxiella burnetii, the causative agent of Q fever, secretes bacterial effector proteins via its Type 4 secretion system to generate a Coxiella-containing vacuole (CCV). Manipulation of lipid and protein trafficking by these effectors is essential for bacterial replication and virulence. Here, we have characterized the lipid composition of CCVs and discovered that the effector Vice interacts with phosphoinositides and membranes enriched in phosphatidylserine (PS) and lysobisphosphatidic acid (LBPA). Remarkably, eukaryotic cells ectopically expressing Vice present compartments that resemble early CCVs in both morphology and composition. We discovered that the biogenesis of these compartments relies on the double function of Vice. The effector protein initially localizes at the plasma membrane of eukaryotic cells where it triggers the internalization of large vacuoles by macropinocytosis. Then, Vice stabilizes these compartments by perturbing the ESCRT machinery and inhibiting the formation of intraluminal vesicles (ILVs). Collectively, our results reveal that Vice is an essential C. burnetii effector protein capable of hijacking two major cellular pathways to shape the bacterial replicative niche. Significance statementCoxiella burnetii is a unique bacterial pathogen that secretes more than a hundred effector proteins to manipulate cellular processes and establish a replicative niche, the Coxiella-containing vacuole (CCV). Our study identified host cell lipids that are actively recruited by the bacterium to the CCV. Using a library of effector mutants, we identified the protein Vice (for Vacuole-inducing Coxiella effector) as the first bacterial effector capable of interacting with lysobisphosphatydic acid-enriched membranes and accumulating this lipid to CCVs. We show that Vice is also capable of stimulating macropinocytosis and inhibiting the ESCRT machinery. Together, our data show how a single bacterial effector can manipulate different cellular processes to favor the biogenesis of a bacterial pathogens niche.

microbiology↗

Reducing stomatal density by expression of a synthetic EPF increases leaf intrinsic water use efficiency and reduces plant water use in a C4 crop

Enhancing crop water use efficiency (WUE) is a key target trait for climatic resilience and expanding cultivation on marginal lands. Reducing stomatal conductance (gs) through manipulating stomatal density has been observed to translate to improved WUE in multiple C3 crop species. However, reducing gs in C3 species often reduces photosynthetic carbon gain. A different response is expected in C4 plants because they possess specialized anatomy and biochemistry which concentrates CO2 at the site of fixation. This modifies the photosynthesis (AN) relationship with intracellular CO2 concentration (ci) so that photosynthesis is CO2-saturated and reductions in gs are unlikely to impair AN. To test this hypothesis, genetic strategies were investigated to reduce stomatal density in the C4 crop sorghum. Constitutive expression of a synthetic epidermal patterning factor (EPF) transgenic allele in sorghum, lead to reduced stomatal densities. A moderate reduction in stomatal density did not strengthen stomatal limitation to AN, improved WUE, reduced water use, and avoided loss of carbon fixation during a period of water deprivation. However, these positive outcomes were associated with negative pleiotropic effects on reproductive development and photosynthetic capacity. Avoiding pleiotropy by targeting expression of the transgene to specific tissues provides a potential pathway to optimal agronomic outcomes.

plant biology↗

Mapping of transgenic alleles in plants using a Nanopore-based sequencing strategy

Transgenic technology was developed to introduce transgenes into various organisms to validate gene function and add genetic variation for the development of beneficial input or output trait over 40 years ago. However, the identification of the transgene insertion position in the genome, while doable, can be cumbersome in the organisms with complex genomes. Here, we report a Nanopore-based sequencing method to rapidly map transgenic alleles in the soybean genome. This strategy is high-throughput, convenient, reliable, and cost-efficient. The transgenic allele mapping protocol outlined herein can be easily translated to other higher eukaryotes with complex genomes.

plant biology↗