Phenotype | EPFL Graph Search

In genetics, the phenotype () is the set of observable characteristics or traits of an organism. The term covers the organism's morphology (physical form and structure), its developmental processes, its biochemical and physiological properties, its behavior, and the products of behavior. An organism's phenotype results from two basic factors: the expression of an organism's genetic code (its genotype) and the influence of environmental factors. Both factors may interact, further affecting the phenotype. When two or more clearly different phenotypes exist in the same population of a species, the species is called polymorphic. A well-documented example of polymorphism is Labrador Retriever coloring; while the coat color depends on many genes, it is clearly seen in the environment as yellow, black, and brown. Richard Dawkins in 1978 and then again in his 1982 book The Extended Phenotype suggested that one can regard bird nests and other built structures such as caddisfly larva cases and beaver dams as "extended phenotypes". Wilhelm Johannsen proposed the genotype–phenotype distinction in 1911 to make clear the difference between an organism's hereditary material and what that hereditary material produces. The distinction resembles that proposed by August Weismann (1834–1914), who distinguished between germ plasm (heredity) and somatic cells (the body). More recently, in The Selfish Gene (1976), Dawkins distinguished these concepts as replicators and vehicles. Despite its seemingly straightforward definition, the concept of the phenotype has hidden subtleties. It may seem that anything dependent on the genotype is a phenotype, including molecules such as RNA and proteins. Most molecules and structures coded by the genetic material are not visible in the appearance of an organism, yet they are observable (for example by Western blotting) and are thus part of the phenotype; human blood groups are an example. It may seem that this goes beyond the original intentions of the concept with its focus on the (living) organism in itself.

How Vibrio cholerae adapts to the environment: from cell shape transitions to antagonistic behavior

Leonardo Filipe Lemos Rocha

The bacterium Vibrio cholerae is the causative agent of the diarrheal disease cholera, which affects millions of people every year. Apart from being a human pathogen, V. cholerae is also a common member of aquatic habitats. Whilst the mechanism that allow it to transit from an environmental to a pathogenic lifestyle is still understudied, it is well established that virulence induction is linked to the bacterium's ability to sense its surroundings. Therefore, this thesis aimed to shed new light on the general sensing capacity of V. cholerae and its adaptation to the surrounding environment. Specifically, we explored the functions of the virulence-associated regulatory two-component system VarS/VarA, which was shown to regulate the major virulence factors of V. cholerae. In this thesis, we identified a new function of this regulatory system in modulating the pathogen's cellular shape. Precisely, we demonstrated that a strain lacking the response regulator VarA loses its normal curve-shaped morphology and transits to spherical shape during stationary phase of growth. This rounding phenotype was not observed on cells lacking the sensor histidine kinase VarS, which suggests the existence of additional VarA activators. Through analysis of the cells's peptidoglycan (PG), we found that the cause of the atypical shape was due to high abundance of dipeptides and reduced peptide cross-links. This abnormal PG composition and consequent cell rounding could be abrogated through cell wall recycling of wild type-derived PG subunits. Through a transposon mutagenesis screen, we identified the carbon storage regulator A (CsrA) as a key player in this phenotype. We showed that varA-deficient cells have an increased activity of CsrA, which led to an overproduction of the aspartate ammonia lyase enzyme (AspA). Our results indicate that AspA overproduction likely reduced the pool of aspartate in varA-deficient cells, which subsequently impaired the biosynthesis of the cell wall precursor meso-diaminopimelic acid. Ultimately, this compromised production of PG precursors resulted in the altered PG composition and consequent cell rounding. Furthermore, and consistent with findings in Enterobacteriaceae, our results also suggest that the VarA-CsrA regulatory circuit modulates chemotaxis and motility in V. cholerae. Finally, in two different collaborative studies, we also addressed how V. cholerae might interact with other bacteria in the environment taking bacterial antagonism strategies into consideration. Firstly, we showed that a subset of human gut microbiota isolates is able to withstand type VI secretion system (T6SS)-mediated attacks by V. cholerae in an immunity protein-independent manner. Moreover, our preliminary data indicates a superior T6SS-mediated killing potential from a specific environmental strain of V. cholerae compared to a particular toxigenic strain. Secondly, we demonstrated that the environmental V. cholerae strain SA7G harbors a specific plasmid, pSA7G1, that confers a competitive advantage over neighboring bacteria likely through type IV secretion system-mediated killing. Collectively, the findings of this thesis contribute to better understanding how V. cholerae adapts to the environment, namely with specific sensing-response mechanisms such as the VarA-CsrA signaling pathway.

EPFL2022

New approaches to uncover the minimally functional hematopoietic niche using the adipocytic differentiation axis: pharmacological modulation and adrenal niche induction

Frédérica Schyrr

Hematopoietic Stem and Progenitor Cells (HSPCs) reside in their niche, a structure that regulates the balance of cellular quiescence, self-renewal and commitment towards differentiated cells. This highly plastic niche is formed by several cellular players, mainly endothelial cells, osteoblasts, adipocytes, and a variety of stromal progenitor cells of which CXCL12-abundant reticular (CAR) cells have been best characterized. Adipocytes have been shown to inhibit hematopoietic progenitor proliferation, but pre-adipocytes can efficiently support the growth of hematopoietic cells in culture. This suggests that the same cellular lineage contains populations with divergent hematopoietic-supportive capacity.In the first part of this thesis, we hypothesized that pharmacological modulation of the adipocytic differentiation axis could inhibit bone marrow (BM)-derived adipocyte maturation and improve hematopoietic progenitor support. Using a phenotypic screen, we identified calcipotriol, a vitamin D derivative, as a strong inhibitor of adipocyte differentiation. In vivo administration of calcipotriol decreased BM adipocyte size at day 18 post BM transplantation, increased total number of colony-forming units (CFUs) per leg and improved the survival of Cxcl12 haploinsufficient mice. Thus, we identified this vitamin D derivative as our top candidate to accelerate hematopoietic recovery in radio/chemotherapy-induced aplasia by targeting the maturation of adipocytes in the BM.Then, we took advantage of a prospective cohort of patients to assess the association between bone health and hematopoietic output in the context of osteoporosis. Osteoporosis is characterized by reduced bone mineral density (BMD), microarchitectural deterioration and increased BM adipocytic content. We studied the association between BMD and differential blood counts. We found that significant associations between blood counts and BMD exist, but none was consistent across bone parameters or timepoints. This suggests that, for steady-state hematopoiesis, there is no clinically significant effect of the deteriorated bone microenvironment on blood production.In the last part, to further investigate the composition of the HSPC-supportive microenvironment, we explored the simpler, albeit functional, extramedullary niche. We showed that the adrenal gland can be transformed into a hematopoietic supportive environment and used as a model to study the minimal components of a de novo niche. This induced adrenal niche supports HSPCs, including serially transplantable hematopoietic stem cells. Circulating HSPCs home into the adrenal cortex, where they are in proximity to CAR cells. These findings are supported by the presence of CXCL12+ cells in human adrenal myelolipoma, a benign tumor that contains extramedullary hematopoietic tissue. We envision our model as a novel tool to study the minimal components needed for hematopoietic support.Thus, using a combination of clinical, translational and basic research, we discovered a new therapeutic candidate approach to improve hematopoietic support by using a vitamin D analog to modify the BM adiposity. We showed the resilience of the BM niche by observing no measurable effect of the osteoporotic niche on hematopoietic output in a large cohort of patients. Finally, we developed a model of de novo hematopoietic niche in the adrenal gland. Altogether, our work provides new strategies to improve our understanding of the hematopoietic niche.

EPFL2022

High-content, targeted RNA-seq screening in organoids for drug discovery in colorectal cancer

Maxim Norkin, Paloma Ordonez Moran

Organoids allow the recapitulation of intestinal homeostasis and cancerogenesis in vitro; however, RNA sequencing (RNA-seq)-based methods for drug screens are missing. We develop targeted organoid sequencing (TORNADO-seq), a high-throughput, high-content drug discovery platform that uses targeted RNA-seq to monitor the expression of large gene signatures for the detailed evaluation of cellular phenotypes in organoids. TORNADO-seq is a fast, highly reproducible time- and cost-effective ($5 per sample) method that can probe cell mixtures and their differentiation state in the intestinal system. We apply this method to isolate drugs that enrich for differentiated cell phenotypes and show that these drugs are highly efficacious against cancer compared to wild-type organoids. Furthermore, TORNADO-seq facilitates in-depth insight into the mode of action of these drugs. Our technology can easily be adapted to many other systems and will allow for more systematic, large-scale, and quantitative approaches to study the biology of complex cellular systems.

2021

New approaches to uncover the minimally functional hematopoietic niche using the adipocytic differentiation axis: pharmacological modulation and adrenal niche induction

Frédérica Schyrr

EPFL2022

How Vibrio cholerae adapts to the environment: from cell shape transitions to antagonistic behavior

Leonardo Filipe Lemos Rocha

EPFL2022