Keith J Micoli

The New York Academy of Sciences, New York, New York, United States

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Publications (8)31.14 Total impact

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    John P Williams · Keith Micoli · Jay M McDonald
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    ABSTRACT: Calcium signaling plays a key role in bone turnover, regulating both osteoblasts and osteoclasts. Despite this the role of calmodulin, the primary intracellular calcium receptor regulatory protein, has received little attention. In this brief review, the function of Ca(2+)/calmodulin signaling in osteoclast development, function, and apoptosis is reviewed. Considerable evidence supports an important regulatory role for Ca(2+)/calmodulin signaling in each of these processes. The overall role of Ca(2+)/calmodulin in regulating bone turnover is also supported by animal and human studies showing that calmodulin antagonists preserve bone mass.
    Preview · Article · Mar 2010 · Annals of the New York Academy of Sciences
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    ABSTRACT: We and others have demonstrated that Fas-mediated apoptosis is a potential therapeutic target for cholangiocarcinoma. Previously, we reported that CaM (calmodulin) antagonists induced apoptosis in cholangiocarcinoma cells through Fas-related mechanisms. Further, we identified a direct interaction between CaM and Fas with recruitment of CaM into the Fas-mediated DISC (death-inducing signalling complex), suggesting a novel role for CaM in Fas signalling. Therefore we characterized the interaction of CaM with proteins recruited into the Fas-mediated DISC, including FADD (Fas-associated death domain)-containing protein, caspase 8 and c-FLIP {cellular FLICE [FADD (Fas-associated death domain)-like interleukin 1beta-converting enzyme]-like inhibitory protein}. A Ca(2+)-dependent direct interaction between CaM and FLIP(L), but not FADD or caspase 8, was demonstrated. Furthermore, a 37.3+/-5.7% increase (n=6, P=0.001) in CaM-FLIP binding was observed at 30 min after Fas stimulation, which returned to the baseline after 60 min and correlated with a Fas-induced increase in intracellular Ca(2+) that reached a peak at 30 min and decreased gradually over 60 min in cholangiocarcinoma cells. A CaM antagonist, TFP (trifluoperazine), inhibited the Fas-induced increase in CaM-FLIP binding concurrent with inhibition of ERK (extracellular-signal-regulated kinase) phosphorylation, a downstream signal of FLIP. Direct binding between CaM and FLIP(L) was demonstrated using recombinant proteins, and a CaM-binding region was identified in amino acids 197-213 of FLIP(L). Compared with overexpression of wild-type FLIP(L) that resulted in decreased spontaneous as well as Fas-induced apoptosis, mutant FLIP(L) with deletion of the CaM-binding region resulted in increased spontaneous and Fas-induced apoptosis in cholangiocarcinoma cells. Understanding the biology of CaM-FLIP binding may provide new therapeutic targets for cholangiocarcinoma and possibly other cancers.
    No preview · Article · Jul 2008 · Biochemical Journal
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    ABSTRACT: The cytosolic domain of human immunodeficiency virus gp160 glycoprotein contains two calmodulin-binding regions. The role of these domains in modulating intracellular calmodulin signaling is of considerable interest in unraveling the mechanism whereby calmodulin regulates Fas-mediated apoptosis in HIV-infected cells. In this investigation we have employed 2D-NMR spectroscopy to determine the solution structure of the 30-residue calmodulin-binding domain corresponding to residues 826-855 of gp160. In solution, the gp160 (826-855) peptide exhibits a high degree of segmental flexibility. Within its conformational manifold, we have detected two separate flexible amphipathic helices involving residues 826-841 and 846-855 connected by a highly flexible type-II beta-turn at Pro-843 and Arg-844. The observed NOE pattern as well as the observation of long-range NOE contacts between the side chains of His-841 and Ile-846 are compatible with the presence of this turn in the conformational manifold of this peptide. This investigation focusing on the properties of the free peptide in solution paves the way for extending the investigations on the interaction of calmodulin with HIV-1 gp160.
    No preview · Article · May 2008 · AIDS research and human retroviruses
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    ABSTRACT: One hallmark of AIDS progression is a decline in CD4+ T lymphocytes, though the mechanism is poorly defined. There is ample evidence that increased apoptosis is responsible for some, if not all, of the decline. Prior studies have shown that binding of cellular calmodulin to the envelope glycoprotein (Env) of HIV-1 increases sensitivity to fas-mediated apoptosis and that calmodulin antagonists can block this effect. We show that individual mutation of five residues in the C-terminal calmodulin-binding domain of Env is sufficient to significantly reduce fas-mediated apoptosis in transfected cells. The A835W mutation in the cytoplasmic domain of gp41 eliminated co-immunoprecipitation of Env with calmodulin in studies with stably transfected cells. Four point mutations (A835W, A838W, A838I, and I842R) and the corresponding region of HIV-1 HXB2 were cloned into the HIV-1 proviral vector pNL4-3 with no significant effect on viral production or envelope expression, although co-immunoprecipitation of calmodulin and Env was decreased in three of these mutant viruses. Only wild-type envelope-containing virus induced significantly elevated levels of spontaneous apoptosis by day 5 post-infection. Fas-mediated apoptosis levels positively correlated with the degree of calmodulin co-immunoprecipitation, with the lowest apoptosis levels occurring in cells infected with the A835W envelope mutation. While spontaneous apoptosis appears to be at least partially calmodulin-independent, the effects of HIV-1 Env on fas-mediated apoptosis are directly related to calmodulin binding.
    Full-text · Article · Feb 2006 · Virology
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    Eric C Seales · Keith J Micoli · Jay M McDonald
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    ABSTRACT: Increased osteoclastic resorption and subsequent bone loss are common features of many debilitating diseases including osteoporosis, bone metastases, Paget's disease, and rheumatoid arthritis. While rapid progress has been made in elucidating the signaling pathways directing osteoclast differentiation and function, a comprehensive picture is far from complete. Here, we explore the role of the Ca(2+)-activated regulator calmodulin in osteoclastic differentiation, functional bone resorption, and apoptosis. During active bone resorption, calmodulin expression is increased, and calmodulin concentrates at the ruffled border, the organelle utilized for acid transport and bone dissolution. Pharmacologic inhibitors of calmodulin, several of which are already used clinically as anti-cancer and anti-psychotic agents, inhibit osteoclastic acid transport, suggesting their potential as bone-sparing drugs. Recent studies also implicate calmodulin in osteoclast apoptosis through a mechanism involving its direct interaction with the death receptor Fas. During osteoclastogenesis, RANKL-induction stimulates a rise in intracellular Ca2+, which in turn activates calmodulin and its downstream effectors. In particular, the Ca(2+)/calmodulin-dependent phosphatase calcineurin and its targets, the NFAT family of transcription factors, have been posited as the master regulators of osteoclastogenesis. However, recent in vivo and in vitro studies demonstrate that another Ca(2+)/calmodulin-regulated effector protein, CaMKII, is also involved. CaMKII(+/-) mutant mice have reduced osteoclast numbers, and CaMKII antagonists inhibit osteoclastogenesis in vitro. Furthermore, CaMKII is known to activate AP-1 transcription factors, which are also required for RANKL-induced osteoclast gene transcription, and recent findings suggest that CaMKII can down-regulate gp130, a cytokine receptor involved in bone remodeling and implicated in numerous osteo-articular diseases.
    Preview · Article · Jan 2006 · Journal of Cellular Biochemistry
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    Chisu Song · Keith Micoli · Eric Hunter
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    ABSTRACT: Mason-Pfizer monkey virus (M-PMV) encodes a transmembrane glycoprotein with a 38-amino-acid-long cytoplasmic tail. After the release of the immature virus, a viral protease-mediated cleavage of the cytoplasmic tail (CT) results in the loss of 17 amino acids from the carboxy terminus and renders the envelope protein fusion competent. To investigate the role of individual amino acid residues in the CT in fusion, a series of mutations was introduced, and the effects of these mutations on glycoprotein biosynthesis and fusion were examined. Most of the alanine-scanning mutations in the CT had little effect on fusion activity. However, four amino acid substitutions (threonine 4, lysine 7, glutamine 9, and isoleucine 10) resulted in substantially increased fusogenicity, while six (leucine 2, phenylalanine 5, isoleucine 13, lysine 16, proline 17, and glycine 31) resulted in much-reduced fusion. Interestingly, the bulk of these mutations are located upstream of the CT cleavage site in a region that has the potential to form a coiled-coil in the Env trimer. Substitutions at glutamine 9 and isoleucine 10 with alanine had the most dramatic positive effect and resulted in the formation of large syncytia. Taken together, these data demonstrate that individual residues within the cytoplasmic domain of M-PMV Env can modulate, in both a positive and negative manner, biological functions that are associated with the extracellular domains of the glycoprotein complex.
    Full-text · Article · Oct 2005 · Journal of Virology
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    ABSTRACT: Assembly of an infectious retrovirus requires the incorporation of the envelope glycoprotein complex during the process of particle budding. We have recently demonstrated that amino acid substitutions of a tyrosine residue in the cytoplasmic domain block glycoprotein incorporation into budding Mason-Pfizer monkey virus (M-PMV) particles and abrogate infectivity (C. Song, S. R. Dubay, and E. Hunter, J. Virol. 77:5192-5200, 2003). To investigate the contribution of other amino acids in the cytoplasmic domain to the process of glycoprotein incorporation, we introduced alanine-scanning mutations into this region of the transmembrane protein. The effects of the mutations on glycoprotein biosynthesis and function, as well as on virus infectivity, have been examined. Mutation of two cytoplasmic residues, valine 20 and histidine 21, inhibits viral protease-mediated cleavage of the cytoplasmic domain that is observed during virion maturation, but the mutant virions show only moderately reduced infectivity. We also demonstrate that the cytoplasmic domain of the M-PMV contains three amino acid residues that are absolutely essential for incorporation of glycoprotein into virions. In addition to the previously identified tyrosine at residue 22, an isoleucine at position 18 and a leucine at position 25 each mediate the process of incorporation and efficient release of virions. While isoleucine 18 may be involved in direct interactions with immature capsids, antibody uptake studies showed that leucine 25 and tyrosine 22 are part of an efficient internalization signal in the cytoplasmic domain of the M-PMV glycoprotein. These results demonstrate that the cytoplasmic domain of M-PMV Env, in part through its YXXL-mediated endocytosis and intracellular trafficking signals, plays a critical role in the incorporation of glycoprotein into virions.
    Full-text · Article · Oct 2005 · Journal of Virology
  • K J Micoli · G Pan · Y Wu · J P Williams · W J Cook · J M McDonald
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    ABSTRACT: Accelerated apoptosis is one mechanism proposed for the loss of CD4+ T-lymphocytes in human immunodeficiency virus type 1 (HIV-1) infection. The HIV-1 envelope glycoprotein, gp160, contains two C-terminal calmodulin-binding domains. Expression of gp160 in Jurkat T-cells results in increased sensitivity to FAS- and ceramide-mediated apoptosis. The pro-apoptotic effect of gp160 expression is blocked by two calmodulin antagonists, tamoxifen and trifluoperazine. This enhanced apoptosis in response to FAS antibody or C(2)-ceramide is associated with activation of caspase 3, a critical mediator of apoptosis. A point mutation in the C-terminal calmodulin-binding domain of gp160 (alanine 835 to tryptophan, A835W) eliminates gp160-dependent enhanced FAS-mediated apoptosis in transiently transfected cells, as well as in vitro calmodulin binding to a peptide corresponding to the C-terminal calmodulin-binding domain of gp160. Stable Tet-off Jurkat cell lines were developed that inducibly express wild type gp160 or gp160A835W. Increasing expression of wild type gp160, but not gp160A835W, correlates with increased calmodulin levels, increased apoptosis, and caspase 3 activation in response to anti-FAS treatment. The data indicate that gp160-enhanced apoptosis is dependent upon calmodulin up-regulation, involves the activation of caspase 3, and requires calmodulin binding to the C-terminal binding domain of gp160.
    No preview · Article · Feb 2000 · Journal of Biological Chemistry

Publication Stats

143 Citations
31.14 Total Impact Points


  • 2010
    • The New York Academy of Sciences
      New York, New York, United States
  • 2000-2006
    • University of Alabama at Birmingham
      • • Department of Pathology
      • • Department of Microbiology
      Birmingham, Alabama, United States