Deciphering the Circadian Interactome

Plants use an endogenous time keeping mechanism known as the circadian clock to regulate the timing of metabolism, physiology and development. However, the basic composition of the circadian system is unknown and how the clock is connected to other pathways in unclear. We combine affinity purification, mass spectrometry, and genetics to identify, dissect, and define the protein complexes within the clock.

Circadian Regulation of Growth

The circadian clock regulates myriad processes in plants. We are particularly interested in how plants adapt their growth to seasonal variation in temperature and day length. We have identified new molecules that act as conduits between the circadian system and growth pathways. We are determining the function of these key proteins with the aim of improving crop production in response to a changing environment.

Tool development for Plants

In order to determine the molecular mechanisms that underlie key plant-specific processes, we are developing new tools and methods to explore protein function. This includes developing methods for rapid, facile purification of protein complexes and new methods for maintaining epitope-tagged proteins at endogenous levels and in their native context. We also develop inexpensive non-invasive imaging systems to monitor plant movement, bioluminescence and fluorescence during long-term time lapse experiments. We make the protocols, instructions, and scripts available and open-source so that we can leverage the community to improve and extend these when possible.

Publications

Pre-prints

Huang H, Gehan M, Huss SE, Alvarez S, Lizarraga C, Gruebbling EL, Gierer J, Naldrett MJ, Bindbeutel R, Evans BS, Mockler TC, Nusinow DA Cross-Species Complementation Reveals Conserved Functions For EARLY FLOWERING 3 Between Monocots And Dicots bioRxiv 131185; doi:10.1101/131185

2017

Lee CM, Adamchek C, Feke A, Nusinow DA, and Gendron JM Mapping Protein–Protein Interactions Using Affinity Purification and Mass Spectrometry Methods In Molecular Biology 1610, no. M110 (2017): doi:10.1007/978-1-4939-7003-2_15

2016

Huang H and Nusinow DA Tandem Purification of His6-3x FLAG Tagged Proteins for Mass Spectrometry from Arabidopsis. Bio-protocol (2016): doi:10.21769/BioProtoc.2060

Huang H and Nusinow DA Into the Evening: Complex Interactions in the Arabidopsis circadian clock. Trends in Genetics (2016): doi:10.1016/j.tig.2016.08.002

Mutka AM, Fentress SJ, Sher JW, Berry JC, Pretz C, Nusinow DA and Bart R Quantitative, image-based phenotyping methods provide insight into spatial and temporal dimensions of plant disease. Plant Physiology (2016): doi:10.1104/pp.16.00984

Huang H, Alvarez S, and Nusinow DA Data on the Identification of Protein Interactors with the Evening Complex and PCH1 in Arabidopsis Using Tandem Affinity Purification and Mass Spectrometry (TAP–MS) Data in Brief (2016): doi:10.1016/j.dib.2016.05.014

Huang H, Yoo CY, Bindbeutel RK, Goldsworthy J, Tielking A, Alvarez S, Naldrett M, Evans B, Chen M, and Nusinow DA. PCH1 integrates circadian and light-signaling pathways to control photoperiod-responsive growth in Arabidopsis. eLife. (2016) doi:10.7554/eLife.13292.

Huang H, Alvarez S, Bindbeutel RK, Shen Z, Naldrett MJ, Evans BS, Briggs SP, Hicks LM, Kay SA, and Nusinow DA. Identification of evening complex associated proteins in Arabidopsis by affinity purification and mass spectrometry. Molecular & Cellular Proteomics. (2016). doi:10.1074/mcp.M115.054064

2015

Kaiserli E, Paldi K, O’Donnell L, Batalov O, Pedmale UV, Nusinow DA, Kay SA, Chory J. Integration of light and photoperiodic signaling in transcriptional nuclear foci. Developmental Cell (2015) doi:10.1016/j,devcel.2015.10.008

Waadt R, Manalansan B, Rauniyar N, Munemasa S, Booker MA, Brandt B, Waadt C, Nusinow, DA, Kay SA, Kunz HH, Schumacher K, DeLong A, Yates JR, and Schroeder JI. Identification of OST1-Interacting Proteins Reveals Interactions with SnRK2-Type Protein Kinases and with PP2A-Type Protein Phosphatases That Function in ABA Responses. Plant Physiology (2015) doi:10.1104/pp.15.00575

2012

Chow BY, Helfer A, Nusinow DA, Kay SA. ELF3 recruitment to the PRR9 promoter requires other Evening Complex members in the Arabidopsis circadian clock. Plant Signaling and Behavior (2012) doi:10.4161/psb.18766

2011

Nusinow DA, Helfer A, Hamilton EE, King JJ, Imaizumi T, Schultz TF, Farre EM, Kay SA. The ELF4-ELF3-LUX Complex Links the Circadian Clock to Diurnal Control of Hypocotyl Growth. Nature (2011) doi:10.1038/nature10182

Helfer A, Nusinow DA, Chow BY, Gehrke AR, Bulyk ML, Kay SA. LUX ARRHYTHMO Encodes a Nighttime Repressor of Circadian Gene Expression in the Arabidopsis Core Clock. Current Biology doi:10.1016/j.cub.2010.12.021

Dmitri A. Nusinow, Ph.D.

Principal Investigator
meter (at) danforthcenter.org
ORCID ID: 0000-0002-0497-1723

He Huang, Ph.D.

Postdoctoral Fellow
hhuang (at) danforthcenter.org
ORCID ID: 0000-0002-4564-7604

Maggie Wilson, Ph.D

Postdoctoral Fellow
MEWilson (at) danforthcenter.org
ORCID ID: Coming Soon

Rebecca Bindbeutel, M.S.

Senior Research Associate
RBindbeutel (at) danforthcenter.org
ORCID ID: 0000-0001-7994-2073

Maria Sorkin

Washington University Graduate Student
MSorkin (at) danforthcenter.org
ORCID ID: 0000-0002-3897-2636

Andrew Lin

Washington University Graduate Student
ALin (at) danforthcenter.org
ORCID ID: 0000-0002-7877-6285

Mervin Schrock

Laboratory technician
MSchrock (at) danforthcenter.org
ORCID ID: Coming

Lab Alumni

2017

Mayla Ayers, Harris Stowe University, NSF–REU

Abigail Hunt, Holt High School, Independent Intern

2016

David Stroshein, Lawrence Technological University, NSF-REU (Graduate School, U of Arizona)

J'Loan Pittman, Washington University, St. Louis, uSTAR Program

2015

Dhruv Patel, Cornell, NSF-REU (Graduate School, UC Berkeley)

Hannah Lucas, U. of Wisconsin, Madison, NSF-REU (Graduate School, Washington U, St. Louis)

Ali Peet, St. Joseph’s Academy, Independent Intern

WIll Wolfe, John Burroughs High School, Independent Intern

2014

Jessica Goldsworthy, Michigan State University, NSF-REU (Pharm D. Program, Ferris State University)

Tom Liu, Ladue High School, STARS Intern

Allison Tielking, MICDS, Independent Intern

2013

Morgan Clark, Western Illinois University, NSF-REU (Chemist, Covance)

Elle Gruebbeling, Timberland High School, STARS Intern

2012

Savannah Est, Holt High School, Independent Intern