Research Summary

proteomic-signaling copyProteomic technology to study cell signaling

Reversible protein phosphorylation is widely used by cells as a signaling mechanism. It regulates, directly or indirectly, most cellular processes. Protein kinases are the central coordinators of signaling, as they are the enzymes responsible for transferring a phosphate group from ATP to targeted protein substrates. Not surprising, perturbations in the action of kinases are associated with several human pathologies, including cancer. Understanding the molecular basis of kinase action and function is of critical importance for biomedical research. It requires knowledge of the kinase substrates, as well as comprehensive characterization of the dynamics and role of the phosphorylation events. Because many kinases are active in a cell and thousands of proteins are phosphorylated, the study of phosphorylation-mediated signaling pathways is challenging and powerful technologies are needed.

We have developed and applied quantitative mass spectrometry technologies for the phosphorylation analysis of protein complexes [1, 2], and for a global screen for in vivo kinase substrates [3, 4]. We are now expanding the use of these technologies to quantitatively characterize signaling dynamics at a proteome-wide scale.


DNA damage checkpoint signaling

In the presence of genotoxic stress, DNA damage checkpoint kinases (see Fig. 2) play a central role in coordinating an elaborate cellular response that involves processes such as DNA repair, DNA replication, cell cycle control and gene transcription. Understanding how the checkpoint kinases contribute to the maintenance of genomic integrity has important implications for cancer research and may provide the basis of rational treatment. We have identified an extensive network of targets of the DNA damage checkpoint kinases in the model organism S. dna-damage-checkpoint copycerevisiae (budding yeast) [3]. A major challenge now is to understand the dynamic regulation of these targets.

We are using quantitative mass spectrometry to systematically characterize the intra-S-phase checkpoint signaling dynamics in yeast. Our goal is to understand how the checkpoint kinases maintain stability of the DNA replication fork, genomic integrity and cell viability.

1. Smolka, M.B., et al., Dynamic Changes in Protein-Protein Interaction and Protein Phosphorylation Probed with Amine-reactive Isotope Tag. Mol Cell Proteomics, 2005. 4(9): p. 1358-69.
2. Smolka, M.B., et al., An FHA domain-mediated protein interaction network of Rad53 reveals its role in polarized cell growth. J Cell Biol, 2006. 175(5): p. 743-53.
3. Smolka, M.B., et al., Proteome-wide identification of in vivo targets of DNA damage checkpoint kinases. PNAS, 2007. 104(25): p. 10364-9.
4. Albuquerque, C.P.*, Smolka, M. B.* et al., A multidimensional chromatography technology for in-depth phosphoproteome analysis. Mol Cell Proteomics, 2008. (*Joint first authors).


Marcus Smolka, PhD

Marcus Smolka is an Associate Professor in the Department of Molecular Biology and Genetics, and a member of the Weill Institute for Cell and Molecular Biology. He received a Ph. D. in Brazil, at the State University of Campinas and worked as a visiting fellow with Dr. Ruedi Aebersold, at the Institute for Systems Biology (Seattle, WA). In 2003, Dr. Smolka moved to San Diego, CA for a post-doctorate at the Ludwig Institute for Cancer Research, working with DNA damage checkpoint kinases and phosphoproteomics in the laboratory of Dr. Huilin Zhou. He joined Cornell in 2008.

Marcus B. Smolka, Ph.D
Weill Institute for Cell and Molecular Biology
Department of Molecular Biology and Genetics
Cornell University
339 Weill Hall
Ithaca, NY 14853-7202

(607)255-0274 (Office)
(607)255-0174 (Lab)

Selected Publications

J Cell Bio (2017) - TOPBP1/Dpb11 plays a conserved role in homologous recombination DNA repair through the coordinated recruitment of 53BP1/Rad9.. Yi Liu, José Renato Cussiol, Jennie Rae Sims, …, Marcus Bustamante Smolka.
Commentary by the Gasser Lab: A game of musical chairs: Pro- and anti-resection factors compete for TOPBP1 binding after DNA damage.

Chromosoma (2016) - Slx4 scaffolding in homologous recombination and checkpoint control: lessons from yeast. Jose Cussiol, Diego Dibitetto, Achille Pellicioli, and Marcus Smolka. [Epub ahead of print] Review.

J Cell Biol (2016) - TOPBP1 takes RADical command in recombinational DNA repair. Yi Liu and Marcus Smolka.

Mol Cell (2015) - Phosphoproteomics Reveals Distinct Modes of Mec1/ATR Signaling during DNA Replication. Francisco Bastos de Oliveira, Dongsung Kim, José Renato Cussiol, Jishnu Das, Min Cheol Jeong, Lillian Doerfler, Kristina Hildegard Schmidt, Haiyuan Yu, Marcus Smolka.

EMBO Journal (2015) - Dampening DNA damage checkpoint signalling via coordinated BRCT domain interactions. José R Cussiol, Carolyn M Jablonowski, Askar Yimit, Grant W Brown, Marcus B Smolka

EMBO Journal (2015) - Assembly of Slx4 signaling complexes behind DNA replication forks. Attila Balint, TaeHyung Kim, David Gallo, Jose Renato Cussiol, Francisco Bastos de Oliveira, Askar Yimit, Jiongwen Ou, Ryuichiro Nakato, Alexey Gurevich, Katsuhiko Shirahige, Marcus Smolka, Zhaolei Zhang, Grant Brown

Nature (2013) - DNA Repair Scaffolds Dampen Checkpoint Signaling by Counteracting the Rad9 Adaptor. Patrice Ohouo, Yi Liu, Francisco M Bastos de Oliveira and Chu Jian "Frank" Ma and Marcus Smolka.

J Biol Chem.(2013) - Interactome analysis reveals ezrin can adopt multiple conformational states. Raghuvir Viswanatha, Jessica Wayt, Patrice Ohouo, Marcus Smolka, Anthony Bretscher.

Cell Cycle (2012) - The many roads to checkpoint activation. Patrice Ohouo and Marcus Smolka.

J Cell Biol (2012) - Local phosphocycling mediated by LOK/SLK restricts ezrin function to the apical aspect of epithelial cells. Raghuvir Viswanatha, Patrice Ohouo, Marcus Smolka, and Anthony Bretscher.

Cell Cycle (2012) - The checkpoint transcriptional response: Make sure to turn it off once you are satisfied. Marcus Smolka, Francisco Bastos de Oliveira, Michael Harris, and Robertus A M de Bruin.

EMBO Journal (2012) - Linking DNA replication checkpoint to MBF cell-cycle transcription reveals a distinct class of G1/S genes. Francisco Bastos de Oliveira, Michael Harris, Pijus Brazauskas, Robertus A M de Bruin and Marcus Smolka.

EMBO Journal (2012) - DNA replication stress differentially regulates G1/S genes via Rad53-dependent inactivation of Nrm1. Anna Travesa, Dwight Kuo, Robertus AM de Bruin, Tatyana I Kalashnikova, Marisela Guaderrama, Kevin Thai, Aaron Aslanian, Marcus Smolka, John R Yates III, Trey Ideker and Curt Wittenberg.

Cell (2011) - TORC1 regulates endocytosis via Npr1-mediated phosphoinhibition of a ubiquitin ligase adaptor. JA MacGurn, PC Hsu, Marcus Smolka and Scott Emr. 147(5):1104-17.

Cell Cycle (2011) - A touching moment for Smc5/6: from ssDNA binding to repair. Patrice Ohouo, Marcus Smolka, (April 2011) (comment on Roy MA, et al. Cell Cycle).

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