PhD – Brandeis University; Professor
Biochemistry of the Cytoskeleton
Our goal is to analyze factors that stabilize the dimer interface in two proteins that serve as subunits for biologically important polymerization reactions. The first of these isthe tubulin a-ß heterodimer, which forms microtubules that are essential for cell transport and cell division in all eukaryotes. Understanding the stability of the intradimer bond in tubulin subunits is important because this bond is virtually identical to the interdimer bond that stabilizes microtubules and because several important antimitotic drugs act by either stabilizing or destabilizing the intersubunit bond in microtubules. The second of these is Cu,Zn superoxide dismutase 1 (SOD1), which forms protofibrils in the familial form of the neurodegenerative diseases Amyotrophic Lateral Sclerosis (FALS). Understanding the stability of the SOD1 intradimer bond is important because we and others have found that dimer dissociation is the first step in the in vitro formation of protofibrils that appear similar to those found in neurons in patients suffering from FALS.
Current work focuses on the role of protein cofactors in the assembly of the tubulin dimer and how the structure of the tubulin dimer interface influences dimer and microtubule stability and dynamics. Surface plasmon resonance is the key tool for these studies. In studies of SOD1 we are determining how different alleles of the protein that result in FALS influence the kinetics of each of the three steps in SOD1 aggregation; i.e., dimer dissociation, loss of metals and protofilament formation.
ATP Mediates Yeast Cell-Cell Communication
Yeast secrete ATP in response to glucose, a property with previously unknown functional consequence. In this report, we show that extracellular ATP is a signal for growth of surrounding cells. The ATP signaling behavior was uncovered by finding reduced toxicity of an inducible, dominant-lethal form of alpha tubulin (tub1-828) in cells grown at high, compared to low cell density. Reduced cell death at high cell density resulted because the rate of chromosome loss/cell division was lower (18-fold) in a cultures inoculated with a high density (350,000) compared to a low density (5,000) of cells. The sparing effect of growth at high cell density could be replicated by growing together 3440 cells that express tub1-828, with 2.3 E6 cells that do not express the mutant protein. Toxicity was reduced at high cell density apparently because a secreted signal induces growth, so that the mutant protein is rapidly diluted by synthesis of wild-type α-tubulin. Further, fluorescence-activated cell sorting (FACS) analysis after DNA staining showed that the rate of the G1-G2 transition was faster with cells at high density. ATP replaced the need for high cell density for resistance to tub1-828, and stimulated the transition from G1 to G2 in cells at low density. Cells lacking the enzyme nucleoside diphosphate kinase did not respond to nucleotide stimulation of growth during expression of mutant tubulin, suggesting that NDP kinase has a regulatory role in growth stimulation. This newly discovered quorum sensing response in yeast, mediated by ATP, indicates that yeast decision-making is not entirely autonomous.
- ATP Mediates Yeast Cell-Cell Communication 2020
- Redler RL, Wilcox KC, Proctor EA, Fee L, Caplow M, Dokholyan NV. Glutathionylation at Cys-111 induces dissociation of wild type and FALS mutant SOD1 dimers. Biochemistry. 2011 Aug 16;50(32):7057-66. Epub 2011 Jul 18.
- Ubiquitin editing enzyme UCH L1 and microtubule dynamics: Implication in mitosis (2010) Anjali Bheda, Anuradha Gullapalli, Michael Caplow, Joseph S. Pagano and Julia Shackelford Cell Cycle, 9, 980-994
- Callahan B, Nguyen K, Collins A, Valdes K, Caplow M, Crossman DK, Steyn AJ, Eisele L, Derbyshire KM. Conservation of structure and protein-protein interactions mediated by the secreted mycobacterial proteins EsxA, EsxB, and EspA. J Bacteriol. 2010 Jan;192(1):326-35.
- Wilcox KC, Zhou L, Jordon JK, Huang Y, Yu Y, Redler RL, Chen X, Caplow M, Dokholyan NV. Modifications of Superoxide Dismutase (SOD1) in Human Erythrocytes: a possible role in ALS. J Biol Chem. 2009 May 15; 284(20):13940-13947.
- Khare SD, Caplow M, Dokholyan NV. FALS mutations in Cu, Zn superoxide dismutase destabilize the dimer and increase dimer dissociation propensity: a large-scale thermodynamic analysis. Amyloid. 2006 Dec;13(4):226-35.
- Khare SD, Caplow M, Dokholyan NV. The rate and equilibrium constants for a multistep reaction sequence for the aggregation of superoxide dismutase in amyotrophic lateral sclerosis. Proc Natl Acad Sci U S A. 2004 Oct 19;101(42):15094-9.
- Caplow M, Fee L. Concerning the chemical nature of tubulin subunits that cap and stabilize microtubules. Biochemistry. 2003 Feb 25;42(7):2122-6.
- Hunter AW, Caplow M, Coy DL, Hancock WO, Diez S, Wordeman L, Howard J. The kinesin-related protein MCAK is a microtubule depolymerase that forms an ATP-hydrolyzing complex at microtubule ends. Mol Cell. 2003 Feb;11(2):445-57.
- Caplow M, Fee L. Dissociation of the tubulin dimer is extremely slow, thermodynamically very unfavorable, and reversible in the absence of an energy source. Mol Biol Cell. 2002 Jun;13(6):2120-31.