Derek W Abbott
Case Western Reserve University
Project start date: 2008-12-09
Project end date: 2013-11-30
Sponsored Links Excellgen http://Excellgen.com
Grants awarded to Derek W Abbott
IRAK4 Kinase Activity´s Role In Immunodeficiency
Derek W Abbott
Pathologycase Western Reserve University
10900 Euclid Ave
cleveland, Oh 441064919
Grant 1R03AI079766-01 from National Institute Of Allergy And Infectious Diseases IRG: ZRG1
Abstract: Upon pathogen exposure, the innate immune system initiates a cytokine response such that the adaptive immune system is tailored to eradicate that pathogen (4, 11, 24). Defects in the innate immune signaling pathways result in patient susceptibility to infectious disease (4, 11, 24, 26). IRAK4 (IL-1 Receptor Associated Kinase 4) is a key signaling molecule that links extracellular pathogen exposure to cytokine transcription and release, and recently, a series of unrelated children who develop recurrent pyogenic infections have been found to have mutations in the IRAK4 gene (6, 15, 16, 22, 25). The mutations in IRAK4 cause C-terminal truncations and result in either the absence of the kinase domain of the molecule or the absence of the total IRAK4 protein (6, 15, 16, 22, 25). IRAK4 is part of a signaling pathway linking the Toll-like Receptors to the NF?B pathway and the MAP Kinase pathways. Mice genetically deleted for IRAK4 show an inability to respond to a variety of pathogen-associated molecules including LPS, peptidoglycan, viral RNA and bacterial DNA. In addition, while showing a normal TNF response, these mice cannot respond to IL-1 (30-33). Surprisingly, patients with mutated IRAK4 show a subtly different phenotype than that found in the mouse as these patients are predominantly susceptible to recurrent gram-positive pyogenic bacterial infections (6, 15, 16, 22, 25, 26). While IRAK4´s kinase activity plays a clear role in these children´s immunodefiency, the role of IRAK4´s kinase activity in TLR signaling is unclear (13, 14, 18, 19, 27). Conflicting data has been published on the requirement of IRAK4 kinase activity for TLR responses and IRAK4´s in vivo substrates and kinetics of activation are unknown. Given the importance of IRAK4 in mediating innate immunity, and given the fact that children with immunodeficiency caused by an IRAK4 mutation all show a deletion in the kinase domain, understanding the kinetics, spatial localization, and specificity of IRAK4´s kinase activity will be important. It will also be important to determine the features that cause differences in IRAK4´s role in gram-positive and gram-negative infections. The central hypothesis of this grant application is that IRAK4´s kinase activity is responsible for effective signaling responses to gram-positive organisms and that differences in this kinase activity (in regards to activity and spatial localization) may underlie the infectious pathology in children with IRAK4 deficiencies. This grant aims to develop a novel in vivo signaling reporter such that IRAK4´s kinase activity can be studied in vivo. If successful, this pilot grant will lead to insights regarding IRAK4´s role in immunodeficiencies and will generate reagents that will help carry the work forward in animal models. People with immunodeficiency syndromes are highly susceptible to infection with bacteria, fungi and viruses. They do not respond normally to these agents, and their immune systems cannot defend them against these pathogens. A mutation in a gene called IRAK4 gives rise to a newly discovered immunodeficiency syndrome. Patients with mutations in IRAK4 develop numerous recurrent infections to fever-causing bacteria. 43% of these patients die in childhood. It is important to understand the function of IRAK4 so that we can better understand the cause of this immunodeficiency syndrome and ultimately, better treat this immunodeficiency syndrome. This grant aims to develop novel reagents with which to study IRAK4 so that this immunodeficiency syndrome can be better understood
Project start date: 2008-07-15
Project end date: 2010-06-30
1R03AI079766-01 (2008): $78500
Innate Immune Signal Transduction Specificity In Inflammatory Disease
Derek W Abbott
Pathologycase Western Reserve University
Grant 1R01GM086550-01 from National Institute Of General Medical Sciences IRG: ZRG1
Abstract: The innate immune system recognizes and responds to pathogenic organisms. In doing so, this system is responsible for initiating a cytokine response designed to tailor the adaptive immune system to eradicate the offending organism. This process must be tightly regulated as too much activity can lead to inflammatory disease. Because inflammatory diseases are characterized by prolonged innate immune activation and cytokine release, the mechanisms controlling downregulation of the innate immune response are paramount in limiting inflammatory pathology. This grant application aims to study the mechanisms of this downregulation by focusing on the signal transduction mechanisms of NOD2 protein (CARD15 gene) and on NOD2´s role in initiating and maintaining the cytokine response. The NOD2 protein is responsible for a number of inflammatory disorders including Blau Syndrome (a familial granulomatosis disease), a subset of Early Onset Sarcoidosis and for 15-20% of genetic Crohn´s Disease. NOD2 is activated in response to intracellular exposure to both gram-positive and gram-negative bacteria after which it helps to coordinate NF-?B activation and cytokine release through the lysine-63 (K63)-linked polyubiquitination of a novel site (K285) on the IKK scaffolding protein NEMO. We have recently extended this finding to show that the major extracellular innate immune signaling receptors, the Toll-like Receptors (TLRs), also require K285 NEMO ubiquitination to properly signal through NF-?B. This work suggests that regulation of the post-translational modifications on the IKK scaffolding protein, NEMO, helps to coordinate cross-talk between intracellular and extracellular innate immune pathways and also helps to regulate the identity, the amount and the duration of cytokines that are released. These findings also suggest that for NF-?B signaling, multiple innate immune signaling pathways converge on NEMO and that the post-translational modifications on NEMO serve as a rheostat to control NF-?B activity. As such, these post-translational modifications may also be targets for molecules aimed at downregulating the NF-?B response activated by NOD2 and other innate immune signaling pathways. The central hypothesis of this grant is that downregulation of NOD2 and TLR-stimulated NF-?B activation is paramount in avoiding inflammatory pathology. Failure to properly downregulate the NF-?B response and coordinate between alternative (MAP kinase) signaling pathways may underlie the pathophysiology of inflammatory disorders. Study of these pathways of downregulation could lead both to novel insight regarding the pathophysiology of these diseases and to novel druggable target to help treat these diseases. To begin to tackle this important problem, we have generated significant preliminary data. We have identified a novel innate immune-induced phosphorylation site on NEMO that controls NEMO ubiquitination and therefore, controls ultimate NF-?B activation. We have also identified a signaling pathway operating through an unexpected MAP3K which inhibits NEMO ubiquitination and shifts innate immune signaling from NF-?B activity toward p38 activity. The Specific Aims of this grant application aim to determine the biochemical mechanisms by which innate immune-induced NF-?B activity can be downregulated, to determine the function of MEKK4 in dictating signal specificity downstream of innate immune system activation and to determine the role of these signaling pathways in the pathophysiology of inflammatory disease. As humans, we are constantly exposed to bacteria, fungi and viruses, and we must respond to these pathogens so that we do not become infected. After responding to these pathogens, if our immune systems do not deactivate, we develop inflammatory disorders such as asthma, inflammatory bowel disease, multiple sclerosis and atherosclerosis (heart and vascular disease). Inflammatory diseases such as these are a significant cause of morbidity and mortality across a wide range of populations (infants to elderly). Due to the importance of downregulating the inflammatory response, our bodies have developed sophisticated mechanisms to dampen the inflammatory response. This grant application aims to study the mechanisms that dampen this inflammatory response and the mechanisms by which this dampening is faulty in inflammatory disease. This work aims to help determine the causes of inflammatory disease and aims to identify novel targets for pharmaceutical intervention in these debilitating disorders
Project start date: 2008-12-09
Project end date: 2013-11-30
RIP2 Kinase´s Function In Innate Immunity
Derek W Abbott
Medicinecase Western Reserve University
10900 Euclid Ave
cleveland, Oh 44106
Grant 5K08AI053819-05 from National Institute Of Allergy And Infectious Diseases IRG: AITC
Abstract: The human body has evolved mechanisms to detect pathogenic organisms and to initiate an appropriate initial immune response against these organisms. Given the diversity of pathogens, a remarkable feature of these innate immunity-signaling pathways is their ability to tailor a response to a particular pathogen. Innate immune responses are initiated at the cell membrane by a family of receptors collectively termed the Toll-like Receptors (TLRs). Downstream signaling pathways activated by the TLRs include the ERK, JNK, p38 and NF- Kappabeta signaling modules. Activation of these signaling pathways then causes cytokine and chemokine release. There is a remarkable specificity to this immune response such that the cytokines released are tailored to the pathogen destined to be eradicated. Although critical, this specificity in signal transduction is poorly understood. The adapter protein kinase RIP2 may mediate a portion of this specificity. RIP2 has been shown to be involved in the proximal TLR signaling pathway as RIP2-null mice show defects in ERK, JNK, p38 and NF-kappaB signaling and subsequent deficiencies in cytokine release in response to LPS from gram-negative bacteria and PGN from gram-positive bacteria. Although RIP2 contains a serine-threonine kinase domain and it readily autophosphorylates, no in vivo RIP2 substrates have been identified. In preliminary studies, we have began to identify the preferred peptide phosphorylation sequence of RIP2. This initial data was used to help identify the adapter protein, TRAF6, as a phosphorylation substrate of RIP2. The central hypothesis of this grant is that as an essential component of the TLR pathway, RIP2´s kinase activity may underly the specificity seen in innate immune signaling pathways. This grant application aims to define the preferred peptide phosphorylation sites of RIP2, to map the phosphorylation site of TRAF6 and to identify the physiologic significance of RIP2´s phosphorylation of TRAF6
Keywords: biological signal transduction, immune response, phosphorylation, protein kinase, toll like receptor JUN kinase, RNA interference, binding protein, cell membrane, cytokine receptor, enzyme activity, mitogen activated protein kinase, nuclear factor kappa beta, serine threonine protein kinase, tumor necrosis factor alpha SDS polyacrylamide gel electrophoresis, immunoprecipitation, mass spectrometry, polymerase chain reaction, tissue /cell culture, western blotting
Project start date: 2004-04-01
Project end date: 2008-01-31
5K08AI053819-05 (2007): $128520
RIP2 Kinase s Function In Innate Immunity
Derek W Abbott
Brigham And Women s Hospital Research Administration Boston, Ma 02115
Grant 5K08AI053819-03 from National Institute Of Allergy And Infectious Diseases IRG: AITC
Abstract: The human body has evolved mechanisms to detect pathogenic organisms and to initiate an appropriate initial immune response against these organisms. Given the diversity of pathogens, a remarkable feature of these innate immunity-signaling pathways is their ability to tailor a response to a particular pathogen. Innate immune responses are initiated at the cell membrane by a family of receptors collectively termed the Toll-like Receptors (TLRs). Downstream signaling pathways activated by the TLRs include the ERK, JNK, p38 and NF- Kappabeta signaling modules. Activation of these signaling pathways then causes cytokine and chemokine release. There is a remarkable specificity to this immune response such that the cytokines released are tailored to the pathogen destined to be eradicated. Although critical, this specificity in signal transduction is poorly understood. The adapter protein kinase RIP2 may mediate a portion of this specificity. RIP2 has been shown to be involved in the proximal TLR signaling pathway as RIP2-null mice show defects in ERK, JNK, p38 and NF-kappaB signaling and subsequent deficiencies in cytokine release in response to LPS from gram-negative bacteria and PGN from gram-positive bacteria. Although RIP2 contains a serine-threonine kinase domain and it readily autophosphorylates, no in vivo RIP2 substrates have been identified. In preliminary studies, we have began to identify the preferred peptide phosphorylation sequence of RIP2. This initial data was used to help identify the adapter protein, TRAF6, as a phosphorylation substrate of RIP2. The central hypothesis of this grant is that as an essential component of the TLR pathway, RIP2 s kinase activity may underly the specificity seen in innate immune signaling pathways. This grant application aims to define the preferred peptide phosphorylation sites of RIP2, to map the phosphorylation site of TRAF6 and to identify the physiologic significance of RIP2 s phosphorylation of TRAF6.
Keywords: biological signal transduction, immune response, phosphorylation, protein kinase, toll like receptor, JUN kinase, RNA interference, binding protein, cell membrane, cytokine receptor, enzyme activity, mitogen activated protein kinase, nuclear factor kappa beta, serine threonine protein kinase, tumor necrosis factor alpha, SDS polyacrylamide gel electrophoresis, immunoprecipitation, mass spectrometry, polymerase chain reaction, tissue /cell culture, western blotting
Project start date: 2004-04-01
Project end date: 2006-10-31
5K08AI053819-03 (2006): $128520
5K08AI053819-02 (2005): $128520
1K08AI053819-01A1 (2004): $108810
Derek W Abbott
Case Western Reserve University
Project start date: 2008-12-09
Project end date: 2013-11-30