Transformative High Resolution Cryo-Electron Microscopy Jon Lorsch, PhD Director National Institute of General Medical Sciences NIH Working Group: NEI, NIGMS, NCI, NHLBI, NIAID, NIDA, NIDDK, NINDS, ORIP, NIAAA, NIBIB Inter-agency Working Group: NSF, DOE, Beckman
Working Group Members Co-Chairs IC Members Jon Lorsch (NIGMS) Dan Gallahan (NCI) Cathy Lewis and Susan Gregurick John (Randy) Knowlton (NCI) Paul Sieving (NEI) Alison Yao (NIAID) Belinda Seto Rao Rapaka (NIDA) Salvatore Sechi (NIDDK) Working Group Coordinators: Paula Flicker (NIGMS) Mary Ann Wu (NIGMS) Denis Buxton (NHLBI) Jim Deatherage (NIGMS) Manjit Hanspal (NHLBI) Houmam Araj (NEI) Jue Chen (NHLBI) James Luo (NHLBI) OSC Margaret Sutherland (NINDS) Ravi Basavappa Malgorzata Klosek (ORIP) Becky Miller Ellie Murcia
Why Now? New Technological Breakthroughs in Cryo-EM 1) New electron microscopy technology dramatically improves our ability to see biological molecules Old Methods New Methods TRPV1 Ion Channel: Mediates burn sensation, Yifan Cheng UCSF 2) New motion correction methods resolve blurring of images due to movement of particles in electron beam Rotavirus Particles Niko Grigorieff, Janelia Farms 3
The U.S. is Rapidly Falling Behind Europe and Asia in Cryo-EM Infrastructure Initial Investment, 1-2 CryoEM microscopes, local facility Significant Investment, 3-4 CryoEM microscopes, regional facility Major Investment, 5+ CryoEM microscopes, HTP comprehensive facility Planned investment
Request for Information (RFI): Transformative High-Resolution Cryo-Electron Microscopy Notice Number: NOT-RM-16-022 Release Date: June 24, 2016 Response Date: August 8, 2016 , 46 responses Training and Workforce Technology Development Need and Capacity • • • HTP pipeline for thin Expertise in all stages Overwhelming support sections of cells required for success for National Centers for automated high- • • Optics and 2 levels of trainees: resolution cryoEM data instrumentation Structural biologists collection for SPA Neophyte biologists • Segmentation and • US is falling behind • subtomogram Hands-on training at • averaging Centers essential CryoET expertise, resources also needed • • Data management and Tutorials, online storage needs materials at home #
Ebola virus neutralization by protective human antibodies (9Å) Two human monoclonal antibodies, mAb100 and mAb114 in combination, protect nonhuman Misasi et al. (2016) Science 351 : 1343 primates against all signs of Ebola virus disease.
Zika Virus 3.8Å Sirohi et al. (2016) Science 352 : 467, 7
HSP90-Cdc37-Cdk4 Complex at 3.9Å • HSP90 Interacts with 60% of human kinome. • Cdk4 kinase is trapped and stabilized in the complex. • HSP90 inhibition leads to degradation of kinases, including oncoproteins vSrc, bRafV600E, Her2. • HSP90 inhibitors are undergoing clinical trials as cancer therapeutics. Verba, K.A. et al. (2016) Science , 352 :1542 8
Challenges and Strategies Challenges ● Cost of instrumentation and upkeep ● Access to high performance data collection ● Limited base of expert investigators Strategy ● Open access to state of the art instrumentation ● Build an expert workforce ● Improve and extend technology ● Create economies of scale Comprehensive Centers ● Three centers ● Research assistance and training 20-30 labs/year ● High-throughput data collection services ● Each center 4 microscopes, 7-8 FTEs, $4M TC/year ● R01/R21s to develop new technology & methods Long Term Plan ● High throughput data collection services at three centers, each $2M TC/year ● Interagency WG (NIH, NSF, DOE, Beckman) Glutamate dehydrogenase (1.8A) Merk et al. Cell 165, 1698, 2016
Proposed Budget Comprehensive Centers (3) Total Year 1 Year 2 Year 3 Year 4 Year 5 $66M Equipment (4 microscopes/center) $22M $22M $22M 0 0 $41.9M Operations $4M $6.4M $10.5M $10.5M $10.5M User Training & Service $9.3M $0.6M $1.2M $2.5M $2.5M $2.5M (4 FTEs/center) $117.2M $26.6M $29.6M $35M $13M $13M Research Grants (R21,R01) Total TC Yearly $15M Cryoelectron Tomography R&D $3M $12M Single Particle R&D $2.4M $27M $5.4M All Activities Total Year 1 Year 2 Year 3 Year 4 Year 5 $144.2M $32M $35M $40.4M $18.4M $18.4M Possible jumpstart program with FY2017 funding of Comprehensive Centers? 10
Thank You! Questions?
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