Purification and characterization of CD38/ADP‐ribosyl cyclase from rat lung

Khoo, Keng Meng; Chang, Chan Fong

doi:10.1080/15216549800201892

Cited by 8 publications

(14 citation statements)

References 12 publications

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“…For example, it has been suggested that two extracellular cysteines in CD38 (Cys119 and Cys201 in human CD38 or Cys123 and Cys205 in mouse CD38) could form interdisulfide bonds between CD38 monomers [22]. In agreement with this hypothesis, studies carried out with porcine heart, rat lung and rat hepatocytes showed that under nonreducing conditions CD38 forms dimers, while under reducing conditions CD38 is present in a monomeric form [23][24][25]. On the other hand, Umar et al have shown that CD38 oligomers, expressed by retinoic acid stimulated HL60 cells, are covalently stabilized by transglutaminase, suggesting an alternate biochemical mechanism for the stabilization of covalent CD38 oligomers [26].…”

supporting

confidence: 66%

CD38 is expressed as noncovalently associated homodimers on the surface of murine B lymphocytes

Moreno‐García

Partida-Sánchez

Primack

et al. 2004

European Journal of Biochemistry

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CD38 is a transmembrane glycoprotein that functions as an ectoenzyme and as a receptor. Based on the structural similarity between CD38 and ADP-ribosyl cyclase from Aplysia californica, it was hypothesized that CD38 is expressed as a homodimer on the surface of cells. Indeed, CD38 dimers have been reported, however, the structural requirements for their stabilization on the plasma membrane are unknown. We demonstrate that the majority of CD38 is assembled as noncovalently associated homodimers on the surface of B cells. Analysis of CD38 mutants, expressed in Ba/F3 cells, revealed that truncation of the cytoplasmic region or mutation of a single amino acid within the a1-helix of CD38 decreased the stability of the CD38 homodimers when solubilized in detergent. Cells expressing the unstable CD38 homodimers had diminished expression of CD38 on the plasma membrane and the half-lives of these CD38 mutant proteins on the plasma membrane were significantly reduced. Together, these results show that CD38 is expressed as noncovalently associated homodimers on the surface of murine B cells and suggest that appropriate assembly of CD38 homodimers may play an important role in stabilizing CD38 on the plasma membrane of B cells.Keywords: B lymphocytes; CD38; homodimer stability; NAD + glycohydrolase; protein structure.CD38 is a type II transmembrane ectoenzyme expressed by many cell types [1][2][3]. CD38 plays an important role in calcium signaling as it catalyzes the production of several calcium mobilizing metabolites including adenosine(5¢)- [4,5]. In addition to its role as an enzyme, CD38 can also serve as a receptor on the plasma membrane of leukocytes and lymphocytes. For example, incubation of B lymphocytes with agonistic antibodies to CD38 induces calcium mobilization, protein phosphorylation, proliferation, class switching, rescue from cell death and induction of apoptosis [1,[6][7][8][9][10]. In order to understand the dual receptor and enzyme properties of CD38, a number of structure/function studies have been performed. These studies have been guided by analyses of two CD38 homologues, the cytosolic Aplysia californica ADP-ribosyl cyclase [11,12] and the mammalian GPI-anchored NAD + glycohydrolase, CD157 [13,14].Crystallographic and X-ray diffraction analyses of these two proteins indicated that both proteins form noncovalently associated homodimers [15,16]. Thus, it has been proposed that CD38 is also likely to be expressed as a homodimer on the plasma membrane and, in agreement with this hypothesis, initial reports showed that high molecular mass aggregates of CD38 are formed after incubation of human erythrocytes with NAD + or 2-mercaptoethanol [17]. In addition, it was reported that CD38 formed dimers and oligomers on the membrane of CD38 transfected HeLa cells [18]. It is not clear, however, whether CD38 is always present in a dimeric form on the surface of cells as many groups have reported finding only the monomeric form of CD38 [19][20][21]. Furthermore, it remains to be determined whether CD38 dimers ar...

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supporting

confidence: 66%

CD38 is expressed as noncovalently associated homodimers on the surface of murine B lymphocytes

Moreno‐García

Partida-Sánchez

Primack

et al. 2004

European Journal of Biochemistry

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“…Purification of CD38 from the Nuclear Fraction-The chromatography steps followed the method of Khoo and Chang (21,22). Briefly, the solubilized nuclear extract was subjected to a series of column chromatography, in the order of blue Sepharose CL-6B, hydroxyapatite, copper-iminodiacetic acid-agarose and concanavalin (Con) A-Sepharose columns.…”

Section: Methodsmentioning

confidence: 99%

“…Antibodies-Production and characterization of the polyclonal antibody against rat CD38 has been previously described (21,22). Various other antibodies were obtained from Affinity Bioreagents Inc. (Golden, CO), including monoclonal against type 1 (RyR-1) and type 2 ryanodine receptor (RyR-2) (clone 34-C), monoclonal anti-calnexin antibody (clone AF18), monoclonal anti-Na ϩ -K ϩ -ATPase antibody (clone 9A-5), polyclonal (rabbit) anti-calreticulin, and polyclonal (rabbit) anti-GRP78 (BiP) antibody.…”

Section: Methodsmentioning

confidence: 99%

“…6, A and B). Since the primary antibody was raised against a polypeptide of 87 amino acid residues consisting of the carboxyl-terminal region of rat CD38 (21,22), it is likely that the enzymatic site of CD38 that is located in the carboxyl-terminal region is facing the nucleoplasm whereby it would have access to NAD ϩ located therein. No immunoreactivity was observed on the outer nuclear envelope or the endoplasmic reticulum regions adjacent to it.…”

Section: Isolation Of a Purified Nuclearmentioning

confidence: 99%

“…The polyclonal antibody used has been shown to be specific for rat (21,22) and murine CD38. 2 This was confirmed in controls using CD38 Ϫ/Ϫ mice (strain C57BL/6), which showed no staining when the anti-CD38 antibody was used under similar conditions (data not shown).…”

Section: Isolation Of a Purified Nuclearmentioning

confidence: 99%

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Localization of the Cyclic ADP-ribose-dependent Calcium Signaling Pathway in Hepatocyte Nucleus

Khoo¹,

Han²,

Park³

et al. 2000

Journal of Biological Chemistry

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CD38 is a type II transmembrane glycoprotein found on both hematopoietic and non-hematopoietic cells. It is known for its involvement in the metabolism of cyclic ADP-ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate, two nucleotides with calcium mobilizing activity independent of inositol trisphosphate. It is generally believed that CD38 is an integral protein with ectoenzymatic activities found mainly on the plasma membrane. Here we show that enzymatically active CD38 is present intracellularly on the nuclear envelope of rat hepatocytes. CD38 isolated from rat liver nuclei possessed both ADP-ribosyl cyclase and NADase activity. Immunofluorescence studies on rat liver cryosections and isolated nuclei localized CD38 to the nuclear envelope of hepatocytes. Subcellular localization via immunoelectron microscopy showed that CD38 is located on the inner nuclear envelope. The isolated nuclei sequestered calcium in an ATP-dependent manner. cADPR elicited a rapid calcium release from the loaded nuclei, which was independent of inositol trisphosphate and was inhibited by 8-amino-cADPR, a specific antagonist of cADPR, and ryanodine. However, nicotinic acid adenine dinucleotide phosphate failed to elicit any calcium release from the nuclear calcium stores. The nuclear localization of CD38 shown in this study suggests a novel role of CD38 in intracellular calcium signaling for non-hematopoietic cells.CD38 is a 42-45-kDa type II transmembrane glycoprotein (1) found in various mammalian tissues and cell types and is capable of converting NAD ϩ into cyclic ADP-ribose (cADPR) 1 (2). This ability is due to the ADP-ribosyl cyclase activity found on the extracellular carboxyl domain of the enzyme. The product, cADPR, possesses calcium mobilizing activity independent of inositol 1,4,5-trisphosphate (IP 3 ) (3). Instead, cADPR seems to regulate calcium mobilization through the calcium-inducedcalcium release mechanism (4, 5). In addition to cyclizing NAD ϩ to cADPR, CD38 is also able to use NADP ϩ as a substrate and catalyze the exchange of its nicotinamide group with nicotinic acid to produce NAADP (5), another potent calciummobilizing metabolite.CD38 is generally believed to be an important surface immunoregulatory molecule; its myriad of possible functions include the induction of B and T cell proliferation (6), regulation of the humoral immune response (7), apoptosis (8), tyrosine phosphorylation of various proteins (9), activation of certain kinases (10), and cytokine release (11). CD38 also displays adhesion properties and might possibly mediate a selectin-type adhesion between different blood populations and human vascular endothelial cells via its putative ligand, CD31 (12). In addition to its immuno-functions, CD38 has been shown to play an important role in insulin secretion via the cADPR-dependent calcium signaling pathway (13). Indeed, results suggest the appearance of autoantibodies to CD38 in patients may contribute to the development of noninsulin-dependent diabetes (14).The calcium stores mobilized by c...

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Insights into nuclear localization and dynamic association of CD38 in Raji and K562 cells

Trubiani

Guarnieri

Eleuterio

et al. 2007

J of Cellular Biochemistry

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CD38 is a type II transmembrane glycoprotein found mainly on the plasma membrane involved in the metabolism of cADPR and NAADP, two nucleotides with calcium mobilizing activity independent of inositol trisphosphate. Recent data report the presence of CD38 in different cellular compartments raising new questions about its effective role in cellular metabolism. In rat hepatocyte nuclei, CD38 has been proposed as a responsive to cADPR integral inner membrane protein suggesting that the nuclear envelope may also be an important source of Ca2+ stores. Further reports indicating that CD38 is localized in nuclear compartments in a variety of cell types and tissues including brain, liver, eye, spleen, and bone raise the condition of resolving the question concerning the effective presence of CD38 within the nucleus. Here we report data supporting the presence of CD38 at nuclear level independently of expression of surface CD38. We utilized two different human leukemia cell lines expressing or not expressing CD38 molecule on their cell surface. The morphological and biochemical results including enzymatic activity and proteomic determinations explain the effective nuclear localization of CD38 in human Raji and K562 cells. Since cell nucleus is a complex and highly dynamic environment with many functionally specialized regions, the nuclear localization of specific proteins represents an important mechanism in signal transduction. The presence of CD38 at the interchromatin region whether linked to nuclear scaffold or stored in nuclear structures as micronuclei and Cajal bodies co-localizing with coilin, suggests its involvement in nuclear processes including transcription, replication, repairing and splicing.

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Purification and characterization of CD38/ADP‐ribosyl cyclase from rat lung

Cited by 8 publications

References 12 publications

CD38 is expressed as noncovalently associated homodimers on the surface of murine B lymphocytes

CD38 is expressed as noncovalently associated homodimers on the surface of murine B lymphocytes

Localization of the Cyclic ADP-ribose-dependent Calcium Signaling Pathway in Hepatocyte Nucleus

Insights into nuclear localization and dynamic association of CD38 in Raji and K562 cells

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