My interest in the compartmentalization of signal transduction, which started with identification of signaling endosomes as a postdoctoral fellow with Dr. William Mobley, led to questions about the proteins that associate with receptor tyrosine kinases in different organelles and membrane fractions. This led to acquisition of phosphoproteomic data from neuroblastoma cell lines and membrane fractions including endosomes and lipid rafts. Analyzing the resulting large data set required acquiring new skills with help from collaborators in the fields of pattern recognition, computational biology and bioinformatics. We discovered that mass spectrometry data, being full of holes (missing values), requires special considerations for embedding statistical relationships into reduced-dimension data structures for clustering. Rigorous clustering provides a filter for network representations of a navigable data structure for exploratory data analysis. Applying these techniques to neuroblastoma phosphoproteomic data and lung cancer data including phosphorylation, methylation, and acetylation, allows predictions about signaling pathways  with much greater resolution than previously possible. 

In my graduate work with Dr. Ed Herbert, I cloned chromogranin A, a major component of neuroendocrine dense core secretory granules. An interest in secretion led me to Dr. Regis Kelly and work in which sorting of regulated and constitutive secretory granules was reconstituted in vitro.  Interest in nerve growth factor signaling mechanisms led me to Dr. William Mobley, in whose laboratory an organelle fractionation approach was employed to define signaling endosomes containing activated TrkA, the NGF receptor tyrosine kinase. These publications have been, and still are, highly cited as the genesis of the signaling endosome hypothesis.

This work fueled further studies on the compartmentalization of signal transduction and mechanisms that affect cell fate decisions, including apoptosis and differentiation. We developed high resolution organelle fractionation techniques to examine endosomes, signaling particles that are resistant to detergent and sensitive to salt, and lipid rafts. We discovered that two key effectors for many pathways, AKT and CREB, are cleaved by caspases during apoptosis. We defined the tipping point of commitment to programmed cell death, after which neuroblastoma and pheochromocytoma cells can no longer be rescued by NGF, and separated cells at different stages of apoptosis by their density. High-resolution organelle fractionation methods revealed that different receptors are in different signaling endosomes distinguished by mass and density. We discovered novel NGF-dependent interactions between TrkA, microtubules, and lipid rafts. These cell biological approaches, combined with sophisticated data analysis techniques, positions my laboratory to make unique and valuable contributions towards understanding signaling networks that control cell fate decisions.

Grimes, M., Hall, B., Foltz, L., Levy, T., Rikova, K., Gaiser, J., Cook, W., Smirnova, E., Wheeler, T., Clark, N. R., Lachmann, A., Zhang, B., Hornbeck, P., Ma’ayan, A., and Comb, M. Integration of protein phosphorylation, acetylation, and methylation data sets to outline lung cancer signaling networks. Sci. Signal. 11, eaaq1087, 2018

Fernandez, N.L., Gundersen, G.W., Rahman, A., Grimes M.L, Rikova, K., Hornbeck, P., and Ma’ayan, A., Clustergrammer, a web-based heatmap visualization and analysis tool for high-dimensional biological data. Sci Data. 4, 170151, 2017. doi: 10.1038/sdata.2017.151

Palacios-Moreno, J., Foltz, L., Guo, A., Stokes, M. P., Kuehn, E. D., George, L., Comb, M., and Grimes, M. L. Neuroblastoma Tyrosine Kinase Signaling Networks Involve FYN and LYN in Endosomes and Lipid Rafts. PLoS Comp Biol 11, 2015. e1004130–e1004133.

Shannon P, Grimes ML, Kutlu B, Bot JJ, Galas DJ. RCytoscape: Tools for Exploratory Network AnalysisBMC Bioinformatics: 14:2172013. doi:10.1186/1471-2105-14-217

Xin X, Gfeller D, Cheng J, Tonikian R, Sun L, et al. SH3 interactome conserves general function over specific formMol Syst Biol 9: 652-669, 2013. doi:10.1038/msb.2013.9.

Grimes, M.L., Lee, W.-J., van der Maarten, L., Shannon, P. Wrangling phosphoproteomic data to elucidate cancer signaling pathways.PLoS ONE 8: e52884. doi:10.1371/journal.pone.0052884.t003. 

Pryor S., McCaffrey G., Young L.R., Grimes M.L.  NGF Causes TrkA to Specifically Attract Microtubules to Lipid Rafts. PLoS ONE 7(4):e35163, 2012. doi:10.1371/journal.pone.0035163

Agnihothram, S. S., B. Dancho, K. W. Grant, M. L. Grimes, D. S. Lyles, and J. H. Nunberg. 2009. Assembly of arenavirus envelopeglycoprotein GPC in detergent-soluble membrane microdomainsJ Virol. 83:9890-9900.

McCaffrey, G., Welker, J.,Scott, J., van der Salm, L., and Grimes, M. L. High-resolution fractionation of signaling endosomescontaining different receptorsTraffic 10, 938-950, 2009.

Lin, D.C., Quevedo, C., Brewer, N.E., Testa, J., Grimes, M.L., Miller, F.D., and Kaplan, D.R. (2006). APPL1 associates with TrkA andGIPC1, and is required for NGF-mediated signal transduction. Mol Cell Biol 26, 8928-8941.

MacCormick, M. Moderscheim, T., van der Salm, L.W.M., Moore, A., Clements, S., McCaffrey, G., and Grimes, M.L.  Distinct signallingparticles containing Erk/Mek and B-Raf in PC12 cells. Biochemical J 387:155-164, 2005.

Weible, M.W., Ozsarac, N., Grimes, M.L., and Hendry, I.A.  Comparison of nerve terminal events in vivo effecting retrograde transport of vesicles containing neurotrophins or synaptic vesicle components. J Neurosci Res 750:771-781, 2004.

Grimes, M.L., and Miettinen, H. Receptor tyrosine kinase and G-protein coupled receptor signalling and sorting within endosomes. JNeurochem, 84: 905-918, 2003.

Francois, F. Godinho, M, Dragunow, M., and Grimes, M.L.  A population of PC12 cells that is initiating apoptosis can be rescued by nerve growth factor, Mol Cell Neurosci, 18:347-3622001.

Francois F, Godinho, MJ, and Grimes M. L. Creb is cleaved by caspases in neural cell apoptosis.  FEBS Lett, 486: 281-284, 2000.

Blythe, T. J., Grimes, M. L. and Kitson, K. E.. "The role of retinoid metabolism by alcohol and aldehyde dehydrogenases in differentiation of cultured neuronal cells." Adv Exp Med Biol 463: 199-204, 1999.

Francois, F, and Grimes, M. L. Phosphorylation-dependent Akt cleavage in neural cell in vitro reconstitution of apoptosis. J. Neurochem.73: 1773-1776, 1999.

Grimes, M. L., Beattie, E., and Mobley, W. C. A signaling organelle containing the nerve growth factor-activated receptor tyrosinekinase, TrkA.  Proc. Nat.Acad. Sci. USA 94: 9909-14, 1997.

Beattie, E. C., Zhou, J., Grimes, M. L., Bunnett, N. W., Howe, C. L., and Mobley, W. C. A signaling endosome hypothesis to explainNGF actions: potential implications for neurodegeneration. Cold Spring Harb. Symp. Quant. Biol. 61: 389-406, 1996.

Grimes, M. L., Zhou, J., Beattie, E., Yuen, E.C., Hall, D.E., Valletta, J.S., Topp, K.S., LaVail, J. H., Bunnett, N.W., and Mobley, W.C.Endocytosis of activated TrkA: Evidence that NGF induces formation of Signalling Endosomes. J. Neurosci. 16:7950-7964, 1996. 

Zhou J, Valetta JS, Grimes ML, and Mobley WC.  Regulation of TrkA expression in PC12 cells after NGF exposure. J.Neurochem.  65:1146-1156, 1995.

Grimes M, Zhou J, Li Y, Holtzman D and Mobley WC.  Neurotrophin signaling in the nervous system.  Seminars in TheNeurosciences 5:239-247, 1993.

Longo FM, Holtzman DM, Grimes M, and Mobley WC:  Nerve Growth Factor:  Actions in the Peripheral and Central Nervous System.  In:  Neurotrophic Factors.  Loughlin S, Fallon J (eds.)  Academic Press, New York, pp. 209-256. 1993. 

Grimes M and Kelly RB.  Sorting of chromogranin B into immature secretory granules in pheochromocytoma, PC12 cells.  In:  Proteases and Protease Inhibitors in Alzheimer?s Disease Pathogenesis.  Banner CDB and Nixon RA  (eds.) Ann. NY Acad.Sci. 674:38-52, 1992.

Grimes M and Kelly RB.  Intermediates in the constitutive and regulated secretory pathways released in vitro  from semi-intact cells.  J. Cell Biol. 117: 539-550, 1992.

Iacangelo A, Grimes M, and Eiden LE.  The bovine chromogranin A gene: Structural basis for hormone regulation and generation of biologically active peptides.  Molec. Endocrin. 5: 1651-1660, 1991.

Lloyd RV, Iacangelo A, Eiden LE, Cano M, Jin L and Grimes M.  Chromogranin A and B messenger ribonucleic acids in pituitary and other normal and neoplastic human endocrine tissues.  Lab. Invest. 60:548-56, 1989.

Grimes M, Iacangelo A, Eiden LE, Godfrey B and Herbert E.  Chromogranin A: the primary structure deduced from cDNA clones reveals the presence of pairs of basic amino acids.  Ann. NY Acad. Sci.  493:351-78, 1987.

Fricker LD,  Liston D, Grimes M and Herbert E.  Specificity of Prohormone Processing:  The Promise of Molecular Biology.  In: Molecular Neurobiology: Recombinant DNA Approaches.  Heinemann S and Patrick J. (eds.) Current Topics in Neurobiology Series, New York: Plenum Press, 1987.

Iacangelo A, Affolter HU, Eiden LE, Herbert E and Grimes M.  Bovine chromogranin A sequence and distribution of its messenger RNA in endocrine tissues.  Nature 323:82-6, 1986.

Nickoloff BJ,  Grimes M,  Wohlfeil E and  Hudson RA.  Affinity-dependent cross-linking to neurotoxin sites of the acetylcholine receptor mediated by catechol oxidation.  Biochemistry 24:999-1007, 1985.

Nickoloff BJ,  Grimes M,  Kelly R and Hudson RA.  Affinity directed reactions of 3-trimethylaminomethyl catechol with the acetylcholine receptor from Torpedo californica.  Biochem. Biophys. Res. Comm. 107:1265-72, 1982.

Tyrosine signaling phosphopeptides from neuroblastoma