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J. Marie Hardwick
J. Marie Hardwick
Molecular Microbiology & Immunology, SPH
Pharmacology and Molecular Sciences
Office (410) 955-2716
Lab (410) 614-3110
Bloomberg School of Public Health
615 N. Wolfe Street, Room E5140
Baltimore, MD 21205
Molecular mechanisms of programmed cell death and its role in viral pathogenesis
Our laboratory studies the basic molecular mechanisms of programmed cell death, an evolutionarily conserved process to eliminate cells. Because these pathways normally contribute to the millions of cell deaths that occur per day per individual, defects in cell death underlie the range of human disorders from cancer (insufficient cell death) to neurological diseases (excessive death). We study these processes in the nervous system, in cancer models and during virus infection using mouse models, yeast genetics and biochemical approaches. We have shown that viruses can trigger cells to activate programmed cell death (Levine et al., Nature, 1993), and that both viral and cellular regulators of apoptosis, such as Bcl-2 family proteins and many other factors, can alter the outcome of a virus infection (Lewis et al., Nature Med, 1999). We seek the mechanisms that explain why Sindbis virus induces neuronal cell death in young animals, but fails to activate the death pathway in neurons of adult brains and in mosquitoes that transmit the virus in nature. Interestingly, cell death factors can also be cell survival factors (Cheng et al., Science, 1997), leading us to pursue their important alternative functions, or ‘day-jobs’, such as regulating mitochondrial fission and bioenergetics, synaptic activity in neurons, or nutrient-sensing in yeast. Although it remains controversial as to whether or not unicellular organisms are capable of undergoing programmed cell death, we have new compelling evidence that these pathways are evolutionarily conserved. Therefore, we have launched an effort to apply the genetic and proteomic tools available for yeast to model the death and survival mechanisms that are conserved between yeast and mammals.
Host cell death responses to virus infections of the brain and of yeast determine disease pathogenesis.
The overall theme under investigation in our lab is the host cell death machinery involved in disease pathogenesis, which started with these early studies of viral persistence, pathogenesis and toxicity. Ongoing studies extend from these studies by characterizing novel biochemical functions of cell death regulators.
Levine B, Huang Q, Isaacs JT, Reed JC, Griffin DE, Hardwick JM, 1993. Conversion of lytic to persistent alphavirus infection by the bcl-2 cellular oncogene. Nature 361, 739-42. https://www.ncbi.nlm.nih.gov/pubmed/8441470 Citations: 519
Lewis J, Oyler GA, Ueno K, Fannjiang YR, Chau BN, Vornov J, Korsmeyer SJ, Zou S, Hardwick JM, 1999. Inhibition of virus-induced neuronal apoptosis by Bax. Nat Med 5, 832-5. https://www.ncbi.nlm.nih.gov/pubmed/10395331 Cited by 103
Ivanovska I, Hardwick JM, 2005. Viruses activate a genetically conserved cell death pathway in a unicellular organism. J Cell Biol 170, 391-9. https://www.ncbi.nlm.nih.gov/pubmed/16061692 Cited by 80. Featured in Editor’s Choice (same issue); Reviewed in AAAS Science STKE 296: p287
Caspase proteases convert Bcl-2 family members and IAP proteins, but not their viral homologs, into potent cell death factors during infection and ischemic brain injury.
Ongoing studies focus on how this conversion to killer mode is regulated in the mouse thymus.
Cheng EH, Kirsch DG, Clem RJ, Ravi R, Kastan MB, Bedi A, Ueno K, Hardwick JM, 1997. Conversion of Bcl-2 to a Bax-like death effector by caspases. Science 278, 1966-8. https://www.ncbi.nlm.nih.gov/pubmed/9395403 Cited by 1297
Clem RJ, Cheng EH, Karp CL, Kirsch DG, Ueno K, Takahashi A, Kastan MB, Griffin DE, Earnshaw WC, Veliuona MA, Hardwick JM, 1998. Modulation of cell death by Bcl-XL through caspase interaction. Proc Natl Acad Sci U S A 95, 554-9. Cited by 532 https://www.ncbi.nlm.nih.gov/pubmed/9435230
Kirsch DG, Doseff A, Chau BN, Lim DS, de Souza-Pinto NC, Hansford R, Kastan MB, Lazebnik YA, Hardwick JM, 1999. Caspase-3-dependent cleavage of Bcl-2 promotes release of cytochrome c. J Biol Chem 274, 21155-61. https://www.ncbi.nlm.nih.gov/pubmed/10409669 Cited by 487
Bellows DS, Chau BN, Lee P, Lazebnik Y, Burns WH, Hardwick JM, 2000. Antiapoptotic herpesvirus Bcl-2 homologs escape caspase-mediated conversion to proapoptotic proteins. J Virol 74, 5024-31. https://www.ncbi.nlm.nih.gov/pubmed/10799576 Cited by 147
Clem RJ, Sheu TT, Richter BW, He WW, Thornberry NA, Duckett CS, Hardwick JM, 2001. c-IAP1 is cleaved by caspases to produce a proapoptotic C-terminal fragment. J Biol Chem 276, 7602-8. https://www.ncbi.nlm.nih.gov/pubmed/11106668 Cited by 123
Seo SY, Chen YB, Ivanovska I, Ranger AM, Hong SJ, Dawson VL, Korsmeyer SJ, Bellows DS, Fannjiang Y, Hardwick JM, 2004. BAD is a pro-survival factor prior to activation of its pro-apoptotic function. J Biol Chem 279, 42240-9. https://www.ncbi.nlm.nih.gov/pubmed/15231831 Cited by 54
Ofengeim D, Chen YB, Miyawaki T, Li H, Sacchetti S, Flannery RJ, Alavian KN, Pontarelli F, Roelofs BA, Hickman JA, Hardwick* JM, Zukin* RS, Jonas* EA, 2012. N-terminally cleaved Bcl-xL mediates ischemia-induced neuronal death. Nat Neurosci 15, 574-80. https://www.ncbi.nlm.nih.gov/pubmed/22366758 Featured in the Scientist, and Neurosci News & Views: Chemo for stroke (same issue); Cited by 38
Non-apoptotic functions of cell death factors in healthy cells.
Before Bcl-2 family proteins and caspases engage the apoptosis pathway they have novel non-apoptotic roles in healthy cells (e.g. regulating neuronal activity, mitochondrial energetics, and more). Ongoing studies investigate novel membrane functions of viral and cellular apoptosis proteins.
Cheng EH, Levine B, Boise LH, Thompson CB, Hardwick JM, 1996. Bax-independent inhibition of apoptosis by Bcl-XL. Nature 379, 554-6. https://www.ncbi.nlm.nih.gov/pubmed/8596636 Cited by 510
Fannjiang Y, Kim CH, Huganir RL, Zou S, Lindsten T, Thompson CB, Mito T, Traystman RJ, Larsen T, Griffin DE, Mandir AS, Dawson TM, Dike S, Sappington AL, Kerr DA, Jonas EA, Kaczmarek LK, Hardwick JM, 2003. BAK alters neuronal excitability and can switch from anti- to pro-death function during postnatal development. Dev Cell 4, 575-85. https://www.ncbi.nlm.nih.gov/pubmed/12689595 Cited by 92
Jonas EA, Hoit D, Hickman JA, Brandt TA, Polster BM, Fannjiang Y, McCarthy E, Montanez MK, Hardwick JM, Kaczmarek LK, 2003. Modulation of synaptic transmission by the BCL-2 family protein BCL-xL. J Neurosci 23, 8423-31. https://www.ncbi.nlm.nih.gov/pubmed/12968005 Featured in: This Week in the Journal (same issue); Cited by 91
Hickman JA, Hardwick JM, Kaczmarek LK, Jonas EA, 2008. Bcl-xL inhibitor ABT-737 reveals a dual role for Bcl-xL in synaptic transmission. J Neurophysiol 99, 1515-22. https://www.ncbi.nlm.nih.gov/pubmed/18160428 Cited by 35.
Berman SB, Chen YB, Qi B, McCaffery JM, Rucker EB, 3rd, Goebbels S, Nave KA, Arnold BA, Jonas EA, Pineda FJ, Hardwick JM, 2009. Bcl-x L increases mitochondrial fission, fusion, and biomass in neurons. J Cell Biol 184, 707-19. https://www.ncbi.nlm.nih.gov/pubmed/19255249; Featured on issue cover; Cited by 150
Alavian KN, Li H, Collis L, Bonanni L, Zeng L, Sacchetti S, Lazrove E, Nabili P, Flaherty B, Graham M, Chen Y, Messerli SM, Mariggio MA, Rahner C, McNay E, Shore GC, Smith PJ, Hardwick JM, Jonas EA, 2011. Bcl-xL regulates metabolic efficiency of neurons through interaction with the mitochondrial F1FO ATP synthase. Nat Cell Biol 13, 1224-33. https://www.ncbi.nlm.nih.gov/pubmed/21926988 Cited by 134
Chen YB, Aon MA, Hsu YT, Soane L, Teng X, McCaffery JM, Cheng WC, Qi B, Li H, Alavian KN, Dayhoff-Brannigan M, Zou S, Pineda FJ, O'Rourke B, Ko YH, Pedersen PL, Kaczmarek LK, Jonas EA, Hardwick JM, 2011. Bcl-xL regulates mitochondrial energetics by stabilizing the inner membrane potential. J Cell Biol 195, 263-76. https://www.ncbi.nlm.nih.gov/pubmed/21987637 Cited by 100
Aouacheria A, Combet C, Tompa P, Hardwick JM, 2015. Redefining the BH3 Death Domain as a 'Short Linear Motif'. Trends Biochem Sci 40, 736-48. https://www.ncbi.nlm.nih.gov/pubmed/26541461 Cited by 12
White K, Arama E, Hardwick JM, 2017. Controlling caspase activity in life and death. PLoS Genet 13, e1006545. https://www.ncbi.nlm.nih.gov/pubmed/28207784
Cell death model system for yeast uncovers prevalence of non-random genome plasticity – one mutation leads to two mutations, and the same pairs of mutant genes co-occur in human tumors.
Using new tools to study gene-dependent cell death in Saccharomyces cerevisiae, we uncovered many surprises, including a widespread phenomenon of directional gene mutation-drive genome evolution, potentially reflective of early steps towards cancer. Ongoing studies seek to translate these findings to mammalian tumorigenesis. Parallel studies in yeast seek to identify yet unknown pathways.
Fannjiang Y, Cheng WC, Lee SJ, Qi B, Pevsner J, McCaffery JM, Hill RB, Basanez G, Hardwick JM, 2004. Mitochondrial fission proteins regulate programmed cell death in yeast. Genes Dev 18, 2785-97.https://www.ncbi.nlm.nih.gov/pubmed/15520274 Cited by 265.
Teng X, Cheng WC, Qi B, Yu TX, Ramachandran K, Boersma MD, Hattier T, Lehmann PV, Pineda FJ, Hardwick JM, 2011. Gene-dependent cell death in yeast. Cell Death Dis 2, e188. https://www.ncbi.nlm.nih.gov/pubmed/21814286 Cited by 25
Teng X, Dayhoff-Brannigan M, Cheng WC, Gilbert CE, Sing CN, Diny NL, Wheelan SJ, Dunham MJ, Boeke JD, Pineda FJ, Hardwick JM, 2013. Genome-wide consequences of deleting any single gene. Mol Cell 52, 485-94. Cited by 53.
https://www.ncbi.nlm.nih.gov/pubmed/24211263 Featured in The Scientist, “One gene, two mutations”; SGD New & Noteworthy, “Gene knockouts may not be so clean after all”; NIH/ NIGMS Biomedical Beat blog by stabilizing the inner membrane potential. J Cell Biol 195, 263-76. https://www.ncbi.nlm.nih.gov/pubmed/21987637 Cited by 100
Yeast cell death genetics – a path to understanding brain function and neurodegeneration.
The top hit in our genome-wide yeast screens was a previously unrecognized homolog of newly identified but uncharacterized disease genes associated with epilepsy and specific cancers. Current studies seek to map nutrient-sensing pathways in mouse epilepsy models.
Cheng WC, Teng X, Park HK, Tucker CM, Dunham MJ, Hardwick JM, 2008. Fis1 deficiency selects for compensatory mutations responsible for cell death and growth control defects. Cell Death Differ 15, 1838-46. https://www.ncbi.nlm.nih.gov/pubmed/18756280 Cited by 38
Hartman AL, Zheng X, Bergbower E, Kennedy M, Hardwick JM, 2010. Seizure tests distinguish intermittent fasting from the ketogenic diet. Epilepsia 51, 1395-402. https://www.ncbi.nlm.nih.gov/pubmed/20477852 Cited by 28
Hartman AL, Santos P, Dolce A, Hardwick JM, 2012. The mTOR inhibitor rapamycin has limited acute anticonvulsant effects in mice. PLoS One 7, e45156. https://www.ncbi.nlm.nih.gov/pubmed/22984623 Cited by 33
Hartman AL, Santos P, O'Riordan KJ, Stafstrom CE, Hardwick JM, 2015. Potent anti-seizure effects of D-leucine. Neurobiol Dis 82, 46-53. https://www.ncbi.nlm.nih.gov/pubmed/26054437 Cited by 7.
Widespread basal (day-job) caspase activity in healthy cells.
Our early work on caspases in regulating viral pathogenesis uncovered clues that caspases have additional non-apoptotic roles in healthy cells prior to activation of cell death. An ultrasensitive caspase biosensor for Drosophila engineered by Hogan Tang, designated CaspaseTracker to study “anastasis”, also provides the first clear evidence of widespread caspase activity in healthy long-lived cells of many fly tissues, including neurons in the brain. Now we seek the functions of these “healthy” caspases.
Nava VE, Rosen A, Veliuona MA, Clem RJ, Levine B, Hardwick JM, 1998. Sindbis virus induces apoptosis through a caspase-dependent, CrmA-sensitive pathway. J Virol 72, 452-9. https://www.ncbi.nlm.nih.gov/pubmed/9420245 Cited by 126
Tang HL, Tang HM, Fung MC, Hardwick JM, 2015. In vivo CaspaseTracker biosensor system for detecting anastasis and non-apoptotic caspase activity. Sci Rep 5, 9015. https://www.ncbi.nlm.nih.gov/pubmed/25757939 Cited by 16.
Tang HL, Tang HM, Fung MC, Hardwick JM, 2016. In Vivo Biosensor Tracks Non-apoptotic Caspase Activity in Drosophila. JoVE https://www.ncbi.nlm.nih.gov/pubmed/27929458