Products Of Separate Genes Research Articles

Kidney in Hypertension

Regulation of blood pressure is a complex integrated response involving a variety of organ systems including the central nervous system (CNS), cardiovascular system, kidneys, and adrenal glands. These systems modulate cardiac output, fluid volumes, and peripheral vascular resistance, the key determinants of blood pressure. More than 40 years ago, Guyton and Coleman1 developed computer models of arterial pressure control, attempting to incorporate the known variables impacting blood pressure homeostasis. The conclusion of this analysis was that regulation of sodium excretion by the kidney and consequent effects on body fluid volumes made up the critical pathway determining the chronic level of intra-arterial pressure. Our own studies of the physiology of blood pressure regulation have focused on the renin-angiotensin (Ang) system (RAS) using genetically modified mouse models. Highly conserved through phylogeny, the RAS is an essential regulator of blood pressure and fluid balance. This biological system is a multienzymatic cascade in which angiotensinogen, its major substrate, is processed in a 2-step reaction by renin and Ang-converting enzyme (ACE), resulting in the sequential generation of Ang I and Ang II. Along with its importance in maintaining normal circulatory homeostasis, abnormal activation of the RAS can contribute to the development of hypertension and target organ damage. The importance of the RAS in clinical medicine is highlighted by the impressive efficacy of pharmacological agents that inhibit the synthesis or activity of Ang II.2–5 At the cellular level, responsiveness to Ang II is conferred by expression of Ang receptors. Ang receptors can be divided into 2 pharmacological classes: type 1 (AT1) and type 2, based on their differential affinities for various nonpeptide antagonists.6,7 Studies using these antagonists suggested that most of the classically recognized functions of the RAS are mediated by AT1 receptors. Gene targeting studies confirmed these conclusions.8 AT …

Identification and characterization of a novel family of membrane magnesium transporters, MMgT1 and MMgT2

Magnesium is an essential metal, but few selective transporters have been identified at the molecular level. Microarray analysis was used to identify two similar transcripts that are upregulated with low extracellular Mg(2+). The corresponding cDNAs encode proteins of 131 and 123 amino acids with two predicted transmembrane domains. The two separate gene products comprise the family that we have termed "membrane Mg(2+) transporters" (MMgTs), because the proteins reside in the membrane and mediate Mg(2+) transport. When expressed in Xenopus laevis oocytes, MMgT1 and MMgT2 mediate Mg(2+) transport as determined with two-electrode voltage-clamp analysis and fluorescence measurements. Transport is saturable Mg(2+) uptake with Michaelis constants of 1.47 +/- 0.17 and 0.58 +/- 0.07 mM, respectively. Real-time RT-PCR demonstrated that MMgT mRNAs are present in a wide variety of cells. Subcellular localization with immunohistochemistry determined that the MMgT1-hemagglutinin (HA) and MMgT2-V5 fusion proteins reside in the Golgi complex and post-Golgi vesicles, including the early endosomes in COS-7 cells transfected with the respective tagged constructs. Interestingly, MMgT1-HA and MMgT2-V5 were found in separate populations of post-Golgi vesicles. MMgT1 and MMgT2 mRNA increased by about threefold, respectively, in kidney epithelial cells cultured in low-magnesium media relative to normal media and in the kidney cortex of mice maintained on low-magnesium diets compared with those animals consuming normal diets. With the increase in transcripts, there was an apparent increase in MMgT1 and MMgT2 protein in the Golgi and post-Golgi vesicles. These experiments suggest that MMgT proteins may provide regulated pathways for Mg(2+) transport in the Golgi and post-Golgi organelles of epithelium-derived cells.

Focus on TGF-β signalling

Transforming growth factor-b (TGF-b) is the index member of a large family of multifunctional growth factors. They were named originally for their ability to induce normal rat kidney fibroblasts to grow in soft agar in the presence of Epidermal Growth Factor (EGF) (De Larco & Todaro 1978, Roberts et al. 1981). There are 5 subtypes, all products of separate genes, three of which, TGF-b1, b2 and b3, have been shown to be expressed in mammalian ovarian cells (Derynck et al. 1988, Hernandez et al. 1990, Mulheron & Schomberg 1990). These factors are synthesized in inactive precursor forms that undergo cleavage to produce monomers, which can dimerize to 25 kDa forms through the conserved cysteine regions. The active regions of the TGF-b monomers share 98–100% identity. The isolation and cloning of inhibin and erythroid differentiation factor (subsequently called activin), and Mullerian inhibiting substance (MIS) led to the observation that these peptides are also members of the TGF-b superfamily. The family has grown to consist of more than 40 members (for a review see Chang et al. 2002), including the growth differentiation factors (GDFs) and bonemorphogenetic proteins (BMPs) (Fig. 1).Members of the family are related by sequence homology (30–40%) and conservation of the cysteine residues. Many also share a similar signal transduction mechanism involving serine–threonine kinase receptors with downstream Smad signalling pathways (for reviews see Miyazawa et al. 2002, Shi & Massague 2003, Lin et al. 2006). The concept that TGF-b superfamily members function as local regulators, either paracrine or autocrine, gained focus after a review on the topic was published by Sporn and Todaro (1980). Further to this, it had been realized that ‘there are events of gametogenesis and endocrine function .which do not necessarily have a direct relationship to the known hormonal milieu being determined by pituitary secretion rates and the afferent blood supply’ (Findlay & Risbridger 1987), supporting the concept of local regulation. It very quickly became apparent that many family members were acting as local regulators in reproductive tissues. In this issue of Reproduction, TGF-b signalling in reproductive tissues is reviewed. This focus issue begins with a chapter on the signal transduction systems that

PlyC: A multimeric bacteriophage lysin

Lysins are murein hydrolases produced by bacteriophage that act on the bacterial host cell wall to release progeny phage. When added extrinsically in their purified form, these enzymes produce total lysis of susceptible Gram-positive bacteria within seconds, suggesting a unique antimicrobial strategy. All known Gram-positive lysins are produced as a single polypeptide containing a catalytic activity domain, which cleaves one of the four major peptidoglycan bonds, and a cell-wall-binding domain, which may bind a species-specific carbohydrate epitope in the cell wall. Here, we have cloned and expressed a unique lysin from the streptococcal bacteriophage C(1), termed PlyC. Molecular characterization of the plyC operon reveals that PlyC is, surprisingly, composed of two separate gene products, PlyCA and PlyCB. Based on biochemical and biophysical studies, the catalytically active PlyC holoenzyme is composed of eight PlyCB subunits for each PlyCA. Inhibitor studies predicted the presence of an active-site cysteine, and bioinformatic analysis revealed a cysteine, histidine-dependent amidohydrolase/peptidase domain within PlyCA. Point mutagenesis confirmed that PlyCA is responsible for the observed catalytic activity, and Cys-333 and His-420 are the active-site residues. PlyCB was found to self-assemble into an octamer, and this complex alone was able to direct streptococcal cell-wall-specific binding. Similar to no other proteins in sequence databases, PlyC defines a previously uncharacterized structural family of cell-wall hydrolases.

Focus on Molecules: Rod photoreceptor cGMP-gated cation channel

The rod photoreceptor cGMP-gated cation (rod CNG) channel is an essential component of phototransduction and also may play a critical role in maintenance of photoreceptor outer segment structural integrity. It is a non-selective cation channel that belongs to the superfamily of voltage-gated channels, and the subfamily of cyclic nucleotide gated channels (Kaupp and Seifert, 2002). This channel is composed of a heterotetramer consisting of three α-subunits (encoded by the CNGA1 locus; also called A subunit) and one β-subunit (encoded by the CNGB1 locus; also called the B subunit; Genbank IDs for Murine and Human mRNA: α-subunit 34328042, 71143140; β-subunit 28386104, 4502918; uniprot IDs for Murine and Human proteins are: α-subunit P29974, P29973; β-subunit Q9UMG2, Q80XL8) arranged around a central pore. The αand β-subunits are the products of separate genes whose loci are designated CNGA1 and CNGB1. The CNGB1 locus is complex encoding multiple alternatively spliced transcripts that are expressed in several diverse tissues (Ardell et al., 2000). For example, an olfactory tissue specific transcript encodes a βsubunit variant that is required for normal channel function of the olfactory cyclic nucleotide gated channel.

Myocyte Enhancer Factors 2A and 2C Induce Dilated Cardiomyopathy in Transgenic Mice

Cardiac hypertrophy and dilation are mediated by neuroendocrine factors and/or mitogens as well as through internal stretch- and stress-sensitive signaling pathways, which in turn transduce alterations in cardiac gene expression through specific signaling pathways. The transcription factor family known as myocyte enhancer factor 2 (MEF2) has been implicated as a signal-responsive mediator of the cardiac transcriptional program. For example, known hypertrophic signaling pathways that utilize calcineurin, calmodulin-dependent protein kinase, and MAPKs can each affect MEF2 activity. Here we demonstrate that MEF2 transcription factors induced dilated cardiomyopathy and lengthening of myocytes. Specifically, multiple transgenic mouse lines with cardiac-specific overexpression of MEF2A or MEF2C presented with cardiomyopathy at base line or were predisposed to more fulminant disease following pressure overload stimulation. The cardiomyopathic response associated with MEF2A and MEF2C was not further altered by activated calcineurin, suggesting that MEF2 functions independently of calcineurin in this response. In cultured cardiomyocytes, MEF2A, MEF2C, and MEF2-VP16 overexpression induced sarcomeric disorganization and focal elongation. Mechanistically, MEF2A and MEF2C each programmed similar profiles of altered gene expression in the heart that included extracellular matrix remodeling, ion handling, and metabolic genes. Indeed, adenoviral transfection of cultured cardiomyocytes with MEF2A or of myocytes from the hearts of MEF2A transgenic adult mice showed reduced transient outward K(+) currents, consistent with the alterations in gene expression observed in transgenic mice and partially suggesting a proximal mechanism underlying MEF2-dependent cardiomyopathy.

Upregulation of the Gamma‐2 Chain of Laminin‐332 (Laminin‐5) in Wounded Mouse Ears

Laminins, extracellular matrix components found in the basement membranes of tissues, are formed from combinations of α, β, and γ polypeptide chains that are separate gene products. Laminin-332 (formerly LN-5) attaches epithelial cells to basement membrane but also affects cell motility during wound healing. In vitro evidence suggests that human acute wound fluid upregulates all three gene products used to assemble laminin-332, but this correlation has not been studied in vivo. Mouse ear skin was wounded by exposure to the chemical warfare agent, bis(2-chloroethyl) sulfide (sulfur mustard, SM) and a time-course study followed to examine the chains of laminin-332 as potential biomarkers of injury. Skin biopsies 24, 72, and 168 hours after SM exposure were analyzed by histology, immunohistochemistry, real-time PCR, and Western blot analysis. SM exposure increased mRNA levels for laminin-γ 2 by 2-fold, 6-fold, and 3-fold for 24, 72, and 168 hours, respectively, but had no observable effect on laminin-α3 or laminin-β3 mRNA. Immunohistochemistry of tissue sections with chain-specific antibodies showed overlap of the three chains extracellularly, around the epithelial cells of the epidermis and those lining the sebaceous glands. Laminin-γ 2 also preferentially accumulated intracellularly. We speculate that the expression of laminin-γ 2 monomers expressed in the SM-treated ears function to enhance migration of the epithelial cells in a manner similar to that reported for invading carcinoma cells. *This work is supported by the USAMRAA under Contract No. DAMD17-02-C-0091 and NIH grant # ES07148.

Highlights from the Literature

Highlights from the Literature

Identification of galectin-4 isoforms in porcine small intestine

Lactose-binding proteins with molecular masses of 14-, 17-, 18-, 28-, and 34-kDa were identified in extracts from porcine small intestinal mucosa. Amino acid sequence analysis of peptides generated by CNBr cleavage of the 34-kDa protein, the most abundant of these proteins, identified this protein as porcine galectin-4. To determine if a porcine homolog of murine galectin-6 is expressed in small intestine, primers for a reverse transcriptase-polymerase chain reaction (RT-PCR) were developed that amplified across the linker region of galectin-4, which is the region that differs between murine galectins-4 and -6. Using these primers, this RT-PCR approach identified two galectin-4 isoforms that differed in the length of their linker region. The larger isoform, galectin-4.1, is nine amino acids longer in its linker region than the smaller isoform, galectin-4.2. Based on nucleotide sequence similarities, the two isoforms are likely splice variants of galectin-4 pre-mRNA and not products of separate genes like murine galectins-4 and -6.

Unexpected similarity in regulation between an archaeal inositol monophosphatase/fructose bisphosphatase and chloroplast fructose bisphosphatase.

Hyperthermophilic archaea have an unusual phosphatase that exhibits activity toward both inositol-1-phosphate and fructose-1,6-bisphosphate, activities carried out by separate gene products in eukaryotes and bacteria. The structures of phosphatases from Archaeoglobus fulgidus (AF2372) and Methanococcus jannaschii (MJ0109), both anaerobic organisms, resemble the dimeric unit of the tetrameric pig kidney fructose bisphosphatase (FBPase). A striking feature of AF2372, but not of MJ0109, is that the sulfhydryl groups of two cysteines, Cys150 and Cys186, are in close proximity (4 A). A similar arrangement of cysteines has been observed in chloroplast FBPases that are regulated by disulfide formation controlled by redox signaling pathways (ferredoxin/thioredoxin). This mode of regulation has not been detected in any other FBPase enzymes. Biochemical assays show that the AF2372 phosphatase activity can be abolished by incubation with O(2). Full activity is restored by incubation with thiol-containing compounds. Neither the C150S variant of AF2372 nor the equivalent phosphatase from M. jannaschii loses activity with oxidation. Oxidation experiments using Escherichia coli thioredoxin, in analogy with the chloroplast FBPase system, indicate an unexpected mode of regulation for AF2372, a key phosphatase in this anaerobic sulfate reducer.

Induction of glutathione synthesis in macrophages by oxidized low-density lipoproteins is mediated by consensus antioxidant response elements.

The uptake of oxidized low-density lipoproteins (oxLDL) by macrophages leading to conversion into foam cells is a seminal event in atherogenesis. Excessive accumulation of oxLDL can cause oxidative stress in foam cells leading to cell death and the progression and destabilization of atherosclerotic lesions. Oxidative stress induces a protective compensatory increase in the synthesis of the endogenous antioxidant glutathione (GSH). Glutamate-cysteine ligase (GCL) is the rate-limiting enzyme in GSH synthesis and is composed of a catalytic subunit (GCLC) and a modifier subunit (GCLM), which are products of separate genes. Treatment of RAW 264.7 mouse macrophages and mouse peritoneal macrophages with oxLDL (30 microg/mL) induces increased expression of both Gclc and Gclm in vitro. The increase in mRNA occurs in part via increased transcription as demonstrated with luciferase reporter constructs. The promoters for both GCLC and GCLM contain consensus antioxidant response elements (AREs). Electrophoretic mobility shift assays revealed induction of nuclear factor binding to these AREs after treatment of RAW 264.7 cells and mouse peritoneal macrophages with oxLDL. Nuclear factor binding to the AREs is diminished by a single base pair substitution in the core sequence. Site-directed mutagenesis of the AREs within the Gclc and Gclm promoters resulted in a decrease of oxLDL-induced luciferase activity. Supershift analyses revealed that oxLDL stimulates binding of the transcription factors Nrf1, Nrf2, and c-jun to the AREs. These data suggest that AREs play a direct role in mediating the induction of GSH synthesis by oxLDL and in protecting macrophages against oxidized lipid-induced oxidative stress.

Two isoforms of Rubisco activase in cotton, the products of separate genes not alternative splicing.

In several plant species, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) activase consists of two isoforms that are produced by alternative splicing of a pre-mRNA. Two forms of activase corresponding to the longer, redox-regulated alpha and the shorter, beta forms were detected immunologically in cotton (Gossypium hirsutum L.) leaves, but their N-termini differed in 4 of 14 residues. The cDNAs for the alpha and beta forms of cotton activase diverged throughout the translated and 3'-untranslated regions, including variations that accounted for the differences in N-terminal amino acid sequence. Analysis of genomic DNA confirmed that separate genes encoded the alpha and beta forms of cotton activase. Separate activase genes were also detected in diploid species of cotton containing the different progenitor genomes of the cultivated allotetraploid, indicating that the occurrence of separate alpha- and beta-form genes in cotton predates the merger of the diploid genomes. The deduced amino acid sequences of the two forms of cotton activase exhibited 84% identity and both forms were active after expression in Escherichia coli. The recombinant alpha and beta forms exhibited similar affinities for ATP and only minor differences in thermotolerance, but their ATPase specific activities differed. The results show for the first time a plant species with two forms of activase that are structurally and functionally equivalent to the alternatively spliced alpha and beta forms in other plants, but that are encoded by separate genes. That cotton still expresses both forms of activase, even without alternative splicing, suggests that each form has a required function in photosynthesis.

The role of thrombospondin-1 in apoptosis.

The thrombospondins are a family of extracellular proteins that participate in cell-to-cell and cell-to-matrix communication. They regulate cellular phenotype during tissue genesis and repair. Five family members, each representing a separate gene product, probably exist in most vertebrate species. Like most extracellular proteins, the thrombospondins are composed of several structural domains that are responsible for the numerous biological functions that have been described for this protein family. Considerable progress has been made towards understanding the function of thrombospondins. The role of thrombospondin in the process of apoptosis or programmed cell death has recently come into focus. In this review we will concentrate on the role of thrombospondin-1 in the broad field of apoptotis research.

Serotonin and norepinephrine transporters: possible relationship between oligomeric structure and channel modes of conduction.

In the past several years there has been significant progress made on the biophysics of neurotransmitter transporters, leading to the proposal of new models of substrate and ion permeation across membranes. Questions arising from these studies are as follows: How are substrate uptake and substrate-induced current related? Where and how does substrate-ion coupling occur? What is the functional significance of the coupled and uncoupled currents? Because of a long-standing interest and collaboration, and because of their importance for normal function and disease, the authors have focused on the properties of human norepinephrine and serotonin transporters, using other clones and mutations as specific needs arise. It has been know for decades that hNETs (human norepinephrine transporters) clear NE+ (norepinephrine) following its release in peripheral sympathetic and central noradrenergic synapses. Neuronal activity influences NE+ uptake, so one is also interested in the acute regulation of hNET. To study these problems, hNET-expressing cells have been developed that are suitable for patch clamp, radioligand uptake, biochemistry, and transiently expressed clones for structure-function analysis, and new protocols have been designed combining patch-clamp, microamperometry, Ca2+ imaging, and native catecholamine transporter preparations to study transporters in whole cells and isolated patches. Using these methods, Na-dependent, NE+-induced hNET currents that are blocked by cocaine and antidepressants, channel modes of NE+ conduction, voltage-dependent uptake coupled to NE+-induced ion channel activity, PKC (phosphokinase C) regulation of NE+ uptake, and transporter modulation by [Ca2+]i have all been discovered. There is also provocative new data on other transporters in this family, such as Li/Na mole fraction experiments in the Drosophila serotonin transporters and sided enkephalin block in proline transporters. These studies have led one to postulate the existence of a narrow pore within transporters through which the substrate (NE+ or serotonin, 5HT+) and other ions (principally Na+) pass. It is hypothesized that the pore resides in an oligomeric structure and that separate gene products of hNET or hSERT (human serotonin transporters) come together to form a channel.

Sexually Dimorphic Expression of Glutamate Decarboxylase mRNA in the Hypothalamus of the Deep Sea Armed Grenadier, Coryphaenoides (Nematonurus) armatus

Glutamate decarboxylase (GAD), is a key enzyme in the central nervous system (CNS) that synthesizes the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) from glutamate. Our previous phylogenetic studies on the evolution of this enzyme indicates that there are at least two distinct forms: GAD65 and GAD67. They are the products of separate genes and probably derive from a common ancestral GAD gene following gene duplication prior to the emergence of the teleosts more than 200 Myr ago. Furthermore, a third GAD-like molecule, GAD3, discovered in the armed grenadier, Coryphaenoides (Nematonurus) armatus, is equally divergent from both GAD65 and GAD67. Specimens of C. (N.) armatus were collected by trawl at a depth of 4,000 m in the Porcupine Seabight (Northeastern Atlantic), and brains dissected and frozen for RNA extraction. All three GAD forms are found in the cerebellum, telencephalon and hypothalamus. Semiquantitative PCR analysis showed that males and females have similar levels of expression of GAD67 and GAD3 in the tissues studied. Independent of the sex examined, the levels of expression of GAD65 and GAD67 in the cerebellum were approximately half that in the telencephalon. GAD3 levels were approximately 30% higher in the cerebellum than in either the telencephalon or hypothalamus. In contrast to GAD67 and GAD3, hypothalamic expression of GAD65 mRNA is 1.8 times higher (p < 0.05) in males than in females. These data indicate that the expression of GAD65, a key enzyme for the synthesis of GABA is sexually dimorphic in females and males of C. (N.) armatus.

Inhibition of focal adhesion kinase prevents growth of prostate tumors in mice

Introduction: Focal adhesion kinase (FAK), a non-receptor mediated tyrosine kinase, is a principal effector regulating adhesion-mediated signals. FAK becomes activated upon integrin-mediated cell attachment and initiates multiple downstream signaling cascades that regulate motility, proliferation, and anchorage-independent growth. Inhibition of adhesion signals blocks these cellular functions and results in cell cycle arrest and the induction of apoptosis. Several studies have shown increased FAK activity in many types of human cancers with the more aggressive tumors expressing larger quantities of activated FAK than less aggressive tumors. In some cells, the carboxy-terminal domain of FAK (called FRNK) is expressed as a separate gene product, and acts as a negative regulator of FAK activity. We hypothesized that unregulated activated FAK facilitates uninhibited tumor growth and that inhibition of these aberrant adhesion signals in prostate cancer cells could prevent tumor implantation and growth in an in vivo mouse model.

Melanoma genetics: An update with focus on the CDKN2A(p16)/ARF tumor suppressors

Investigative interest in atypical nevi and familial melanoma has contributed to the identification of several candidate melanoma loci within the human genome. Molecular defects in both tumor suppressor genes and oncogenes have been pathogenically linked to melanoma in recent studies. Of the loci currently characterized, the major gene resides on chromosome 9p and encodes a tumor suppressor designated p16. This gene, which is also known as CDKN2A, is either mutated or deleted in a large majority of melanoma cell lines, as well as in many uncultured melanoma cells and in the germline of melanoma kindreds. A novel aspect of the p16 locus is that it encodes not just one but two separate gene products that are transcribed in alternative reading frames. Both products function as negative regulators of cell cycle progression. The p16 protein itself executes its effects by competitively inhibiting cyclin-dependent kinase 4, which is a factor necessary for cellular progression through a major regulatory transition of the cell division cycle. Inherited and acquired deletions or point mutations in the p16 gene increase the likelihood that potentially mutagenic DNA damage will escape repair before cell division. Notably, the second product of the locus, ARF (for alternative reading frame), regulates cell growth through independent effects on the p53 pathway. Although there is little evidence that ARF by itself is involved in the pathogenesis of melanoma, deletions at the p16 locus disable two separate pathways that control cell growth. These recent advances open up the possibility of genetic testing for melanoma susceptibility in the setting of familial melanoma and suggest novel therapeutic strategies for melanoma based on gene therapy or small molecule mimicry targeted to the correction of defects in the p16 regulatory pathway. (J Am Acad Dermatol 2000;42:705-22.) Learning Objective: At the conclusion of this learning activity, participants should be familiar with the historical aspects of melanoma genetics and should have a greater understanding of the CDKN2A(p16)/ARF tumor suppressor genes.

Neuropeptide Y family of peptides: Structure, anatomical expression, function, and molecular evolution

Evolutionary relationships between neuroendocrine peptides are often difficult to resolve across divergent phyla due to independent duplication events in different lineages. Thanks to peptide purification and molecular cloning in many different species, the situation is beginning to clear for the neuropeptide Y (NPY) family, which also includes peptide YY (PYY), the tetrapod pancreatic polypeptide (PP) and the fish pancreatic peptide Y (PY). It has long been assumed that the first duplication to occur in vertebrate evolution generated NPY and PYY, as both of these are found in all gnathostomes as well as lamprey. Evidence from other gene families show that this duplication was probably a chromosome duplication event. The origin of a second PYY peptide found in lamprey remains to be explained. Our recent cloning of NPY, PYY and PY in the sea bass proves that fish PY is a separate gene product. We favour the hypothesis that PY is a duplicate of the PYY gene and that it may have occurred late in fish evolution, as PY has so far only been found in acanthomorph fishes. Thus, this duplication seems to be independent of the one that generate PP from PYY in tetrapods, although both tetrapod PP and fish PY are expressed in the pancreas. Studies in the sea bass and other fish show that PY, in contrast to PP, is expressed in the nervous system. We review the literature on the distribution and functional aspects of the various NPY-family peptides in vertebrates.

Cardiovascular α1-adrenoceptor subtypes: functions and signaling

alpha-Adrenoceptors (alpha1AR) are G protein-coupled receptors and include alpha1A, alpha1B, and alpha1D subtypes corresponding to cloned alpha1a, alpha1b, and alpha1d, respectively. alpha1AR mediate several cardiovascular actions of sympathomimetic amines such as vasoconstriction and cardiac inotropy, hypertrophy, metabolism, and remodeling. alpha1AR subtypes are products of separate genes and differ in structure, G protein-coupling, tissue distribution, signaling, regulation, and functions. Both alpha(1A)AR and alpha(1B)AR mediate positive inotropic responses. On the other hand, cardiac hypertrophy is primarily mediated by alpha(1A)AR. The only demonstrated major function of alpha(1D)AR is vasoconstriction. alpha1AR are coupled to phospholipase C, phospholipase D, and phospholipase A2; they increase intracellular Ca2+ and myofibrillar sensitivity to Ca2+ and cause translocation of specific phosphokinase C isoforms to the particulate fraction. Cardiac hypertrophic responses to alpha1AR agonists might involve activation of phosphokinase C and mitogen-activated protein kinase via Gq x alpha1AR subtypes might interact with each other and with other receptors and signaling mechanisms.

Structure and characterization of the genes for murine choline/ethanolamine kinase isozymes α and β

AbstractCholine/ethanolamine kinase (CK/EK) is the first enzyme in phosphatidylcholine/phosphatidylethanolamine biosynthesis in all animal cells. The highly purified CKs from mammalian sources and their recombinant gene products so far were all shown to have EK activity also, indicating that both activities reside on the same protein. CK/EK in most animal cells exists as several isoforms, for two of which (α and β) their cDNAs have been cloned from both the rat and mouse, and they are found to be separate gene products. The physiological significance for the existence of more than one CK/EK enzyme, however, remains to be clarified. In this study, we isolated mouse genes encoding both types of CK/EK isozyme and determined their entire structure. The 5′-flanking promoter regions were found to have quite different features from each other, indicating that their expression could be under distinct control. Comparison of the nucleotide sequence between the corresponding coding exons showed the best homology (75%) residing on exon VIII. A search of the database resulted in the possible existence of 17 different origins of eukaryotic CK and/or EK, each of which presumably contained the entire amino acid sequence. Multialignment of their putative amino acid sequences led to an identification of the novel consensus sequence possibly required for the expression of either CK or EK activity, which corresponded to the sequence within exons VII and VIII of CK/EK-α and -β genes from the mouse. This sequence was localized in close proximity to the C-terminal region of the general (Brenner's) phosphotransferase concensus sequence which was also completely conserved in all of the putative eukaryotic CK/EK proteins. The results demonstrated that, while both CK/EK-α and -β genes were composed of 11 major exons, the size of their genes was quite different: 40 kb for CK/EK-α, whereas it was only 3.5 kb for CK/EK-β. —Aoyama, C., N. Yamazaki, H. Terada, and K. Ishidate. Structure and characterization of the genes for murine choline/ethanolamine kinase isozymes α and β. J. Lipid Res. 2000. 41: 452–464.