Renal epithelial cells must maintain distinct protein compositions in their apical and basolateral membranes in order to perform their transport functions. The creation of these polarized protein distributions depends on sorting signals that designate the trafficking route and site of ultimate functional residence for each protein. Segregation of newly synthesized apical and basolateral proteins into distinct carrier vesicles can occur at the trans-Golgi network, recycling endosomes, or a growing assortment of stations along the cellular trafficking pathway. The nature of the specific sorting signal and the mechanism through which it is interpreted can influence the route a protein takes through the cell. Cell type-specific variations in the targeting motifs of a protein, as are evident for Na,K-ATPase, demonstrate a remarkable capacity to adapt sorting pathways to different developmental states or physiologic requirements. This review summarizes our current understanding of apical and basolateral trafficking routes in polarized epithelial cells.

The ability to switch a gene from off to on and monitor dynamic changes provides a powerful approach for probing gene function and elucidating causal regulatory relationships. Here, we developed and characterized YETI (Yeast Estradiol strains with Titratable Induction), a collection in which > 5,600 yeast genes are engineered for transcriptional inducibility with single‐gene precision at their native loci and without plasmids. Each strain contains SGA screening markers and a unique barcode, enabling high‐throughput genetics. We characterized YETI using growth phenotyping and BAR‐seq screens, and we used a YETI allele to identify the regulon of Rof1, showing that it acts to repress transcription. We observed that strains with inducible essential genes that have low native expression can often grow without inducer. Analysis of data from eukaryotic and prokaryotic systems shows that native expression is a variable that can bias promoter‐perturbing screens, including CRISPRi. We engineered a second expression system, Z 3 EB42, that gives lower expression than Z 3 EV, a feature enabling conditional activation and repression of lowly expressed essential genes that grow without inducer in the YETI library.

Genome-wide phenotypic screens in the budding yeast Saccharomyces cerevisiae have produced the largest, richest and most systematic phenotypic description of any organism. Such an achievement was enabled by the development of highly scalable phenotypic assays and construction of the yeast knock-out (YKO) collection, comprising ~5,000 isogenic strains each deleted for exactly one open reading frame. Systematic screening of the YKO collection led to ~500 publications describing ~14,500 phenotypes capturing nearly every aspect of yeast biology. Yet, integrative analyses of this rich data source have been virtually impossible due to the lack of a central repository and consistent meta-data annotations. Here, we describe the aggregation, harmonization and analysis of all published phenotypic screens of the YKO collection, which we refer to as the Yeast Phenome (www.yeastphenome.org). To demonstrate the power of data integration and illustrate how much it facilitates the generation of testable hypotheses, we present three discoveries uniquely enabled by Yeast Phenome. First, we use the variation in the number of phenotypes per gene to identify tryptophan homeostasis as a central point of vulnerability to a wide range of chemical compounds, including FDA-approved drugs. Second, using phenotypic profiles as a tool for predicting gene function, we identify and validate the role of YHR045W as a novel regulator of ergosterol biosynthesis and DNA damage response, and YGL117W as a new member of the aromatic amino acid biosynthesis pathway. Finally, we describe a surprising exponential relationship between phenotypic similarity and intergenic distance in both yeast and human genomes. This relationship, which stretches as far as 380 kb in yeast and 100 Mb in humans, suggests that gene positions are optimized for function to a much greater extent than appreciated previously. Overall, we show that Yeast Phenome enables systematic enquiries into the nature of gene-gene and gene-phenotype relationships and is an important new resource for systems biology.

In coated vesicle formation, coat protein recruitment needs to be spatially and temporally controlled. The coating process involves conformational changes of the coat protein complexes that activate them for interaction with cargo or machinery components and coat polymerization. Here we discuss mechanisms that have emerged recently from studies of the clathrin adaptor and the COPI systems.