Annual Review of Physiology - Volume 65, 2003

The tubuloglomerular feedback response, the change in afferent arteriolar tone caused by a change in NaCl concentration at the macula densa, is likely initiated by the generation of a vasoactive mediator within the confines of the juxtaglomerular apparatus. Substantial progress has been made in identifying the nature of this mediator and the factors that modulate its effect on vascular tone. In support of earlier studies using P1 purinergic antagonists, the application of the knockout technique has shown that adenosine 1 receptors are absolutely required for eliciting TGF responses. The background level of angiotensin II appears to be an important cofactor determining the efficiency of A1AR-induced vasoconstriction, probably through a synergistic interaction at the level of the G protein–dependent transduction mechanism. The source of the adenosine is still unclear, but it is conceivable that adenosine is generated extracellularly from released ATP through a cascade of ecto-nucleotidases. There is also evidence that ATP may activate P2 receptors in preglomerular vessels, which may contribute to autoregulation of renal vascular resistance. Nitric oxide (NO), generated by the neuronal isoform of nitric oxide synthase in macula densa cells, reduces the constrictor effect of adenosine, but the regulation of NO release and its exact role in states of TGF-induced hyperfiltration are still unclear.

The physiological tuning and pathophysiological alterations of renal proximal reabsorption of inorganic phosphate can be ascribed to the net amount of the Na/Pi-cotransporter NaPi-IIa localized in the brush border membrane. The net amount of NaPi-IIa appears to be the result of an endocytotic rate regulated by a complex network of different protein kinases. New approaches demonstrated that NaPi-IIa is part of heteromeric protein complexes, organized by PDZ (postsynaptic protein PSD95, Drosophila junction protein Disc-large, tight junction protein ZO-1) proteins. Such complexes are thought to play important roles in the apical positioning and regulated endocytosis of NaPi-IIa and therefore such interactions have to be considered when explaining proximal phosphate ion reabsorption.

Urea plays a key role in the urine-concentrating mechanism. Physiologic and molecular data demonstrate that urea transport in kidney and red blood cells occurs by specific urea transporter proteins. Two gene families for facilitated urea transporters, UT-A and UT-B, and several urea transporter cDNA isoforms have been cloned from human, rat, mouse, and several non-mammalian species. Polyclonal antibodies have been generated to many of the urea transporter proteins, and several novel findings have resulted from their use in integrative animal studies. For example, (a) vasopressin increases the phosphorylation of UT-A1 in rat inner medullary collecting duct; (b) UT-A1 protein abundance is increased in the rat inner medulla during conditions in which urine-concentrating ability is reduced; and (c) urea transporters are expressed in non-renal tissues, and UT-A protein abundance is up-regulated in uremia in both liver and heart. In addition to the facilitated urea transporters, functional evidence exists for active urea transport in the kidney collecting duct. This review summarizes the physiologic evidence for the existence of facilitated and active urea transporters, the molecular biology of the facilitated urea transporter gene families and cDNAs, and integrative studies into urea transporter protein regulation, both in the kidney and in other organs.

During the response to metabolic acidosis, the intercalated cell of the collecting tubule converts from one that secretes HCO3⁻ to one that absorbs HCO3⁻ by H⁺ secretion. The molecular basis of this complex change in phenotype was studied in an immortalized intercalated cell line. We found that it was induced by secretion, polymerization, and deposition of a protein, which we termed hensin, into the extracellular matrix. Surprisingly, this change in phenotype is identical to terminal differentiation of epithelial cells in that it recapitulated all the characteristics of terminal differentiation, including a change in cell shape, acquisition of specialized apical structures (microvilli and ruffles), and the ability to secrete and endocytose materials in a regulated manner from the apical membrane. Hensin is expressed in most epithelia, and others have discovered that it is deleted in a large number of epithelial tumors. These results suggest that conversion of polarity of the intercalated cells represents a process of terminal differentiation.

Gene therapy as a treatment modality for pulmonary disorders has attracted significant interest over the past decade. Since the initiation of the first clinical trials for cystic fibrosis lung disease using recombinant adenovirus in the early 1990s, the field has encountered numerous obstacles including vector inflammation, inefficient delivery, and vector production. Despite these obstacles, enthusiasm for lung gene therapy remains high. In part, this enthusiasm is fueled through the diligence of numerous researchers whose studies continue to reveal great potential of new gene transfer vectors that demonstrate increased tropism for airway epithelia. Several newly identified serotypes of adeno-associated virus have demonstrated substantial promise in animal models and will likely surface soon in clinical trials. Furthermore, an increased understanding of vector biology has also led to the development of new technologies to enhance the efficiency and selectivity of gene delivery to the lung. Although the promise of gene therapy to the lung has yet to be realized, the recent concentrated efforts in the field that focus on the basic virology of vector development will undoubtedly reap great rewards over the next decade in treating lung diseases.

A number of conditions, such as pneumonia, trauma, or systemic sepsis arising from the gut, may result in the acute respiratory distress syndrome (ARDS). Because of its significant morbidity and mortality, ARDS has been the focus of extensive research. One specific area of interest has been the investigation of the role of the surfactant system in the pathophysiology of this disease. Several studies have demonstrated that alterations of surfactant contribute to the lung dysfunction associated with ARDS, which has led to investigations into the use of exogenous surfactant as a therapy for this syndrome. Clinical experience with surfactant therapy has been variable owing to a number of factors including the nature of the injury at the time of treatment, the specific surfactant preparation utilized, the dose and delivery method chosen, the timing of surfactant administration over the course of the disease, and the mode of ventilation used during and after surfactant administration.

The alveolar macrophage responds to bacterial infection with the production of inflammatory mediators that include TNFα. Early production of TNFα results in increased bacterial clearance, whereas too much TNFα results in many of the hallmarks of bacterial sepsis. TNFα production is regulated at many levels, including multiple signaling pathways, that lead to transcription, translation, and release of functional TNFα. Interactions of mitogen-activated protein (MAP) kinases, lipid signaling pathways, and oxidant-mediated mechanisms regulate the response of alveolar macrophages to infection. Animal models of sepsis support the central role played by macrophage-derived TNFα in sepsis.

Understanding of the functions and regulation of the phenotype of the alveolar type I epithelial cell has lagged behind studies of its neighbor the type II cell because of lack of cell-specific molecular markers. The recent identification of several proteins expressed by type I cells indicates that these cells may play important roles in regulation of cell proliferation, ion transport and water flow, metabolism of peptides, modulation of macrophage functions, and signaling events in the peripheral lung. Cell systems and reagents are available to characterize type I cell biology in detail, an important goal given that the cells provide the extensive surface that facilitates gas exchange in the intact animal.

Maintenance of membrane lipid asymmetry is a dynamic process that influences many events over the lifespan of the cell. With few exceptions, most cells restrict the bulk of the aminophospholipids to the inner membrane leaflet by means of specific transporters. Working in concert with each other, these proteins correct for sporadic incursions of the aminophospholipids to the outer membrane leaflet as a result of bilayer imbalances created by various cellular events. A shift in the relative contribution in each of these activities can result in sustained exposure of the aminophospholipids at the cell surface, which allows capture of the cells by phagocytes before the integrity of the plasma membrane is compromised. The absence of an efficient recognition and elimination mechanism can result in uncontrolled and persistent presentation of self-antigens to the immune system, with development of autoimmune syndromes. To prevent this, phagocytes have developed a diverse array of distinct and redundant receptor systems that drive the postphagocytic events along pathways that facilitate cross-talk between the homeostatic and the immune systems. In this work, we review the basis for the proposed mechanism(s) by which apoptotic ligands appear on the target cell surface and the phagocyte receptors that recognize these moieties.

In Drosophila photoreceptors, the light-sensitive current is mediated downstream of phospholipase C by TRP (transient receptor potential) channels. Recent evidence suggests that Drosophila TRP channels are activated by diacylglycerol (DAG) or its metabolites (polyunsaturated fatty acids), possibly in combination with the reduction in phosphatidyl inositol 4,5 bisphosphate (PIP2). Consistent with this view, diacylglycerol kinase is identified as a key enzyme required for response termination. Signaling is critically dependent upon efficient PIP2 synthesis; mutants of this pathway in combination with genetically targeted PIP2 reporters provide unique insights into the kinetics and regulation of PIP2 turnover. Recent evidence indicates that a growing number of mammalian TRP homologues are also regulated by lipid messengers, including DAG, arachidonic acid, and PIP2.

Phosphoinositides [PPIs, which collectively refer to phosphorylated derivatives of phosphatidylinositol (PI)] have a pivotal role as precursors to important second messengers and as bona fide signaling and scaffold targeting molecules. This review focuses on recent advances that elucidate how PPIs, particularly PI(4,5)P2 (PIP2), directly regulate the actin cytoskeleton in vivo by modulating the activity and targeting of actin regulatory proteins. The role of PIP2 in stimulating actin polymerization and in establishing cytoskeleton–plasma membrane linkages is emphasized. In addition, the review presents tantalizing evidence that suggests how binding of selected cytoskeletal proteins to membrane PPIs may promote PPI clustering into raft lipid microdomains, alter their accessibility to other proteins, and even distort the bilayer conformation. These actions have profound implications for many other PPI-regulated membrane functions that are beginning to be uncovered, and they suggest how PPIs can mediate crosstalk between the actin cytoskeleton and an expanding spectrum of essential cellular functions.

The phosphoinositides PtdIns(4,5)P2 and PtdIns(3,4,5)P3 are concentrated in plasma membranes of eukaryotic cells, and excluded from endosomes, whereas PtdIns(3)P is formed in these latter intracellular membranes and is apparently excluded from the plasma membrane. The logic of this asymmetric disposition is now revealed by the nature of the effector proteins that selectively bind these lipids through specific modules and by the processes that they catalyze. PtdIns(3,4,5)P3 has a role in directing exocytosis, in addition to many other signaling events, whereas PtdIns(4,5)P2 directs endocytosis through its ability to anchor several coat proteins to the plasma membrane. Remarkably, the elimination of PtdIns(4,5)P2 from forming endosomes may be required for membrane fission to occur. Thus membrane insertion and retrieval can be regulated by plasma membrane concentrations of PtdIns(3,4,5)P3 and PtdIns(4,5)P2, whereas PtdIns(3)P directs the downstream trafficking and recycling of intracellular membranes through its attraction of proteins that catalyze these processes. The phosphoinositides thereby control many cell features that depend upon protein sorting, including the composition of the plasma membrane itself, which in turn determines the cell's responses to its environment.

The cell membrane Na,K-ATPase is a member of the P-type family of active cation transport proteins. Recently the molecular structure of the related sarcoplasmic reticulum Ca-ATPase in an E1 conformation has been determined at 2.6 Å resolution. Furthermore, theoretical models of the Ca-ATPase in E2 conformations are available. As a result of these developments, these structural data have allowed construction of homology models that address the central questions of mechanism of active cation transport by all P-type cation pumps. This review relates recent evidence on functional sites of Na,K-ATPase for the substrate (ATP), the essential cofactor (Mg²⁺ ions), and the transported cations (Na⁺ and K⁺) to the molecular structure. The essential elements of the Ca-ATPase structure, including 10 transmembrane helices and well-defined N, P, and A cytoplasmic domains, are common to all PII-type pumps such as Na,K-ATPase and H,K-ATPases. However, for Na,K-ATPase and H,K-ATPase, which consist of both α- and β-subunits, there may be some detailed differences in regions of subunit interactions. Mutagenesis, proteolytic cleavage, and transition metal-catalyzed oxidative cleavages are providing much evidence about residues involved in binding of Na⁺, K⁺, ATP, and Mg²⁺ ions and changes accompanying E1-E2 or E1-P-E2-P conformational transitions. We discuss this evidence in relation to N, P, and A cytoplasmic domain interactions, and long-range interactions between the active site and the Na⁺ and K⁺ sites in the transmembrane segments, for the different steps of the catalytic cycle.

Rhodopsin is a retinal photoreceptor protein of bipartite structure consisting of the transmembrane protein opsin and a light-sensitive chromophore 11-cis-retinal, linked to opsin via a protonated Schiff base. Studies on rhodopsin have unveiled many structural and functional features that are common to a large and pharmacologically important group of proteins from the G protein-coupled receptor (GPCR) superfamily, of which rhodopsin is the best-studied member. In this work, we focus on structural features of rhodopsin as revealed by many biochemical and structural investigations. In particular, the high-resolution structure of bovine rhodopsin provides a template for understanding how GPCRs work. We describe the sensitivity and complexity of rhodopsin that lead to its important role in vision.