Ift88 is a central component of the intraflagellar transport (Ift) complex B, essential for the building of cilia and flagella from single cell organisms to mammals. Loss of Ift88 results in the absence of cilia and causes left-right asymmetry defects, disordered Hedgehog signaling, and polycystic kidney disease, all of which are explained by aberrant ciliary function. In addition, a number of extraciliary functions of Ift88 have been described that affect the cell-cycle, mitosis, and targeting of the T-cell receptor to the immunological synapse. Similarly, another essential ciliary molecule, the kinesin-2 subunit Kif3a, which transports Ift-B in the cilium, affects microtubule (MT) dynamics at the leading edge of migrating cells independently of cilia. We now show that loss of Ift88 impairs cell migration irrespective of cilia. Ift88 is required for the polarization of migrating MDCK cells, and Ift88 depleted cells have fewer MTs at the leading edge. Neither MT dynamics nor MT nucleation are dependent on Ift88. Our findings dissociate the function of Ift88 from Kif3a outside the cilium and suggest a novel extraciliary function for Ift88. Future studies need to address what unifying mechanism underlies the different extraciliary functions of Ift88.

The primary cilium has evolved as a multifunctional cellular compartment that decorates most vertebrate cells. Cilia sense mechanical stimuli in various organs, but the molecular mechanisms that convert the deflection of cilia into intracellular calcium transients have remained elusive. Polycystin-2 (TRPP2), an ion channel mutated in polycystic kidney disease, is required for cilia-mediated calcium transients but lacks mechanosensitive properties. We find here that TRPP2 utilizes TRPV4 to form a mechano- and thermosensitive molecular sensor in the cilium. Depletion of TRPV4 in renal epithelial cells abolishes flow-induced calcium transients, demonstrating that TRPV4, like TRPP2, is an essential component of the ciliary mechanosensor. Because TRPV4-deficient zebrafish and mice lack renal cysts, our findings challenge the concept that defective ciliary flow sensing constitutes the fundamental mechanism of cystogenesis.

Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations in PKD1 or PKD2, the genes encoding polycystin 1 (PC1) and polycystin 2 (PC2), respectively. PC1 and PC2 localize to the primary cilium and form a protein complex, which is thought to regulate signaling events. PKD1 mutations are associated with a stronger phenotype than PKD2, suggesting the existence of PC1 specific functions in renal tubular cells. However, the evidence for diverging molecular functions is scant. The bending of cilia by fluid flow induces a reduction in cell size through a mechanism that involves the kinase LKB1 but not PC2. Here, using different in vitro approaches, we show that contrary to PC2, PC1 regulates cell size under flow and thus phenocopies the loss of cilia. PC1 is required to couple mechanical deflection of cilia to mTOR in tubular cells. This study pinpoints divergent functions of the polycystins in renal tubular cells that may be relevant to disease severity in ADPKD.

Kidney diseases still pose one of the biggest challenges for global health, and their heterogeneity and often high comorbidity load seriously hinders the unraveling of their underlying pathomechanisms and the delivery of optimal patient care. Metabolomics, the quantitative study of small organic compounds, called metabolites, in a biological specimen, is gaining more and more importance in nephrology research. Conducting a metabolomics study in human kidney disease cohorts, however, requires thorough knowledge about the key workflow steps: study planning, sample collection, metabolomics data acquisition and preprocessing, statistical/bioinformatics data analysis, and results interpretation within a biomedical context. This review provides a guide for future metabolomics studies in human kidney disease cohorts. We will offer an overview of important a priori considerations for metabolomics cohort studies, available analytical as well as statistical/bioinformatics data analysis techniques, and subsequent interpretation of metabolic findings. We will further point out potential research questions for metabolomics studies in the context of kidney diseases and summarize the main results and data availability of important studies already conducted in this field.

The opportunistic bacterium Pseudomonas aeruginosa can infect mucosal tissues of the human body. To persist at the mucosal barrier, this highly adaptable pathogen has evolved many strategies, including invasion of host cells. Here, we show that the P. aeruginosa lectin LecB binds and cross-links fucosylated receptors at the apical plasma membrane of epithelial cells. This triggers a signaling cascade via Src kinases and phosphoinositide 3-kinase (PI3K), leading to the formation of patches enriched with the basolateral marker phosphatidylinositol (3,4,5)-trisphosphate (PIP3) at the apical plasma membrane. This identifies LecB as a causative bacterial factor for activating this well-known host cell response that is elicited upon apical binding of P. aeruginosa. Downstream from PI3K, Rac1 is activated to cause actin rearrangement and the outgrowth of protrusions at the apical plasma membrane. LecB-triggered PI3K activation also results in aberrant recruitment of caveolin-1 to the apical domain. In addition, we reveal a positive feedback loop between PI3K activation and apical caveolin-1 recruitment, which provides a mechanistic explanation for the previously observed implication of caveolin-1 in P. aeruginosa host cell invasion. Interestingly, LecB treatment also reversibly removes primary cilia. To directly prove the role of LecB for bacterial uptake, we coated bacterium-sized beads with LecB, which drastically enhanced their endocytosis. Furthermore, LecB deletion and LecB inhibition with l-fucose diminished the invasion efficiency of P. aeruginosa bacteria. Taken together, the results of our study identify LecB as a missing link that can explain how PI3K signaling and caveolin-1 recruitment are triggered to facilitate invasion of epithelial cells from the apical side by P. aeruginosa. IMPORTANCE An intriguing feature of the bacterium P. aeruginosa is its ability to colonize highly diverse niches. P. aeruginosa can, besides forming biofilms, also enter and proliferate within epithelial host cells. Moreover, research during recent years has shown that P. aeruginosa possesses many different mechanisms to invade host cells. In this study, we identify LecB as a novel invasion factor. In particular, we show that LecB activates PI3K signaling, which is connected via a positive feedback loop to apical caveolin-1 recruitment and leads to actin rearrangement at the apical plasma membrane. This provides a unifying explanation for the previously reported implication of PI3K and caveolin-1 in host cell invasion by P. aeruginosa. In addition, our study adds a further function to the remarkable repertoire of the lectin LecB, which is all brought about by the capability of LecB to recognize fucosylated glycans on many different niche-specific host cell receptors.

Cilia are microtubule-based organelles that assemble via intraflagellar transport (IFT) and function as signaling hubs on eukaryotic cells. IFT relies on molecular motors and IFT complexes that mediate the contacts with ciliary cargo. To elucidate the architecture of the IFT-B complex, we reconstituted and purified the nonameric IFT-B core from Chlamydomonas reinhardtii and determined the crystal structures of C. reinhardtii IFT70/52 and Tetrahymena IFT52/46 subcomplexes. The 2.5-Å resolution IFT70/52 structure shows that IFT52330-370 is buried deeply within the IFT70 tetratricopeptide repeat superhelix. Furthermore, the polycystic kidney disease protein IFT88 binds IFT52281-329 in a complex that interacts directly with IFT70/IFT52330-381 in trans. The structure of IFT52C/IFT46C was solved at 2.3 Å resolution, and we show that it is essential for IFT-B core integrity by mediating interaction between IFT88/70/52/46 and IFT81/74/27/25/22 subcomplexes. Consistent with this, overexpression of mammalian IFT52C in MDCK cells is dominant-negative and causes IFT protein mislocalization and disrupted ciliogenesis. These data further rationalize several ciliogenesis phenotypes of IFT mutant strains.

Chronic kidney disease (CKD) represents a chronic proinflammatory state and is associated with very high cardiovascular risk. Apolipoprotein A-IV (apoA-IV) has antiatherogenic, antioxidative, anti-inflammatory and antithrombotic properties and levels increase significantly during the course of CKD.

The kidneys operate at the interface of plasma and urine by clearing molecular waste products while retaining valuable solutes. Genetic studies of paired plasma and urine metabolomes may identify underlying processes. We conducted genome-wide studies of 1,916 plasma and urine metabolites and detected 1,299 significant associations. Associations with 40% of implicated metabolites would have been missed by studying plasma alone. We detected urine-specific findings that provide information about metabolite reabsorption in the kidney, such as aquaporin (AQP)-7-mediated glycerol transport, and different metabolomic footprints of kidney-expressed proteins in plasma and urine that are consistent with their localization and function, including the transporters NaDC3 (SLC13A3) and ASBT (SLC10A2). Shared genetic determinants of 7,073 metabolite-disease combinations represent a resource to better understand metabolic diseases and revealed connections of dipeptidase 1 with circulating digestive enzymes and with hypertension. Extending genetic studies of the metabolome beyond plasma yields unique insights into processes at the interface of body compartments.

Chronic kidney disease (CKD), a major public health problem with differing disease etiologies, leads to complications, comorbidities, polypharmacy, and mortality. Monitoring disease progression and personalized treatment efforts are crucial for long-term patient outcomes. Physicians need to integrate different data levels, e.g., clinical parameters, biomarkers, and drug information, with medical knowledge. Clinical decision support systems (CDSS) can tackle these issues and improve patient management. Knowledge about the awareness and implementation of CDSS in Germany within the field of nephrology is scarce.

Cilia are specialized organelles that play an important role in several biological processes, including mechanosensation, photoperception, and osmosignaling. Mutations in proteins localized to cilia have been implicated in a growing number of human diseases. In this study, we demonstrate that the von Hippel-Lindau (VHL) protein (pVHL) is a ciliary protein that controls ciliogenesis in kidney cells. Knockdown of pVHL impeded the formation of cilia in mouse inner medullary collecting duct 3 kidney cells, whereas the expression of pVHL in VHL-negative renal cancer cells rescued the ciliogenesis defect. Using green fluorescent protein-tagged end-binding protein 1 to label microtubule plus ends, we found that pVHL does not affect the microtubule growth rate but is needed to orient the growth of microtubules toward the cell periphery, a prerequisite for the formation of cilia. Furthermore, pVHL interacts with the Par3-Par6-atypical PKC complex, suggesting a mechanism for linking polarity pathways to microtubule capture and ciliogenesis.

Omics data facilitate the gain of novel insights into the pathophysiology of diseases and, consequently, their diagnosis, treatment, and prevention. To this end, omics data are integrated with other data types, e.g., clinical, phenotypic, and demographic parameters of categorical or continuous nature. We exemplify this data integration issue for a chronic kidney disease (CKD) study, comprising complex clinical, demographic, and one-dimensional 1H nuclear magnetic resonance metabolic variables. Routine analysis screens for associations of single metabolic features with clinical parameters while accounting for confounders typically chosen by expert knowledge. This knowledge can be incomplete or unavailable. We introduce a framework for data integration that intrinsically adjusts for confounding variables. We give its mathematical and algorithmic foundation, provide a state-of-the-art implementation, and evaluate its performance by sanity checks and predictive performance assessment on independent test data. Particularly, we show that discovered associations remain significant after variable adjustment based on expert knowledge. In contrast, we illustrate that associations discovered in routine univariate screening approaches can be biased by incorrect or incomplete expert knowledge. Our data integration approach reveals important associations between CKD comorbidities and metabolites, including novel associations of the plasma metabolite trimethylamine-N-oxide with cardiac arrhythmia and infarction in CKD stage 3 patients.

The microtubular motor Kinesin-2 and its subunit Kif3a are essential for the formation of primary cilia, an organelle implicated in a wide spectrum of developmental abnormalities. Outside cilia, Kinesin-2 mediated transport has been implicated in vesicle and N-cadherin transport, but it is unknown if and how extraciliary Kif3a affects basic cellular functions such as migration or the formation of multicellular structures. Here we show that tetracycline inducible depletion of Kif3a in MDCK cells slows epithelial cell migration. Microtubules at the leading edge of Kif3a depleted cells failed to grow perpendicularly into the leading edge and microtubular dynamics were dampened in Kif3a depleted cells. Loss of Kif3a retarded lateral membrane specification and completely prevented the formation of three-dimensional spheres in collagen. These data uncover that Kif3a regulates the microtubular cytoskeleton in the cell periphery and imply that extra-ciliary Kif3a has an unexpected function in morphogenesis.

Inactivation of the tumor suppressor von Hippel-Lindau (VHL) gene is a key event in hereditary and sporadic clear cell renal cell carcinomas (ccRCC). The mechanistic target of rapamycin (mTOR) signaling pathway is a fundamental regulator of cell growth and proliferation, and hyperactivation of mTOR signaling is a common finding in VHL-dependent ccRCC. Deregulation of mTOR signaling correlates with tumor progression and poor outcome in patients with ccRCC. Here, we report that the regulatory-associated protein of mTOR (RAPTOR) is strikingly repressed by VHL. VHL interacts with RAPTOR and increases RAPTOR degradation by ubiquitination, thereby inhibiting mTORC1 signaling. Consistent with hyperactivation of mTORC1 signaling in VHL-deficient ccRCC, we observed that loss of vhl-1 function in C. elegans increased mTORC1 activity, supporting an evolutionary conserved mechanism. Our work reveals important new mechanistic insight into deregulation of mTORC1 signaling in ccRCC and links VHL directly to the control of RAPTOR/mTORC1. This may represent a novel mechanism whereby loss of VHL affects organ integrity and tumor behavior.

The primary cilium is a sensory organelle at the cell surface with integral functions in cell signaling. It contains a microtubular axoneme that is rooted in the basal body (BB) and serves as a scaffold for the movement of intraflagellar transport (IFT) particles by Kinesin-2 along the cilium. Ift88, a member of the anterograde moving IFT-B1 complex, as well as the Kinesin-2 subunit Kif3a are required for cilia formation. To facilitate signaling, the cilium restricts the access of molecules to its membrane ("ciliary gate"). This is thought to be mediated by cytoskeletal barriers ("subciliary domains") originating from the BB subdistal/distal appendages, the periciliary membrane compartment (PCMC) as well as the transition fibers and zone (TF/TZ). The PCMC is a poorly characterized membrane domain surrounding the ciliary base with exclusion of certain apical membrane proteins. Here we describe that Ift88, but not Kinesin-2, is required for the establishment of the PCMC in MDCK cells. Likewise, in C. elegans mutants of the Ift88 ortholog osm-5 fail to establish the PCMC, while Kinesin-2 deficient osm-3 mutants form PCMCs normally. Furthermore, disruption of IFT-B1 into two subcomplexes, while disrupting ciliogenesis, does not interfere with PCMC formation. Our findings suggest that cilia are not a prerequisite for the formation of the PCMC, and that separate machineries with partially overlapping functions are required for the establishment of each.