Enter the text here that is the new abstract information for your application. This section must be no longer than 30 lines of text. This proposal focuses on the dynamic behavior of mitochondria in cells. In particular, it examines two pathways that control that behavior via different posttranslational modifications of the motor-adaptor protein milton/TRAK1/2. The movement of mitochondria is driven by two microtubule-based motors, Kinesin-1 and Dynein, and these motors are bound to the mitochondrial surface by a motor/adaptor complex consisting of the proteins milton and Miro (also called TRAK1/2 and RhoT1/2). The first regulatory pathway we here investigate involves the enzyme O-GlcNAc Transferase (OGT), an enzyme that catalyzes the addition of the sugar residue N-Acetyl Glucosamine to serine and threonine residues on proteins. We and others have found that this enzyme GlcNAcylates milton. We also find that expression of OGT in neurons arrests the movement of the mitochondria and that this occurs directly through its substrate milton. Because OGT is thought to be a nutrient sensor that is more active when glucose levels are high, we hypothesized that the OGT pathway will stop mitochondria under conditions of high extracellular glucose and indeed we observe that mitochondrial movement is decreased in axons via this pathway when glucose levels are raised. We propose to examine the mechanism by which OGT halts mitochondria and also the significance of OGT and milton GlcNAcylation for the distribution of mitochondria in axons. The second regulatory pathway that is examined in this grant also concerns mitochondrial/cytoskeletal interactions and their regulation by a posttranslational modification, in this case phosphorylation of milton. Through the motors that move mitochondria and also additional likely anchoring proteins, mitochondria normally exist in a close relationship with microtubules. We find that this situation is very different in dividing cells. When a cell enters mitosis, the microtubules are dismantled and reform as the spindle apparatus. During mitosis, mitochondria are excluded from the regions of the cell that contain microtubules. We have found that milton becomes phosphorylated during the cell cycle, probably by cdk1. We will investigate the significance of this phosphorylation for the state of the motor/adaptor complex and its significance for the mitotic redistribution of mitochondria.

Public Health Relevance

NO CHANGE FROM PREVIOUS) Mitochondria and their dynamics are crucial to the survival of many cell types, but especially neurons. The present proposal examines two pathways regulating their dynamics: one that may help neurons cope with a changing nutrient environment and another that may be crucial during the division of non-neuronal cells. Defects in controlling mitochondrial dynamics are likely to cause neurodegeneration and may also impair proper inheritance of chromosomes by daughter cells. From: Applegate, Theresa To: Urdaneta, Angela (NIH/NIGMS) [E];Gindhart, Joe (NIH/NIGMS) [E] Cc: Schwarz, Thomas Subject: 2R01GM069808 - 09A1 PI Name: Schwarz, Thomas L. (BCH) Date: Friday, August 22, 2014 3:50:09 PM Attachments: 2R01GM06908-09A1 Schwarz Scope.pdf Dear Dr. Gindhardt and Ms. Urdaneta, We received the news that the competing renewal of Dr. Schwarzs Milton and the Transport of Mitochondria project (2R01GM069808 - 09A1) will be awarded, but at a reduced rate of funding. Therefore, we are requesting permission to change the scope of work by removing Specific Aim 1C. Dr. Schwarzs commitment to the project remains unchanged at 1.8 calendar months effort. Attached, please find a copy of Dr. Schwarzs original request along with updated versions of the Project Summary/Abstract and the Specific Aims sections of the application. Should you have any questions or if you need additional information, please do not hesitate to contact me. Thank you for your continued support. Best regards, Theresa Theresa Applegate Boston Childrens Hospital Manager of Sponsored Programs 300 Longwood Avenue Boston, MA 02115 P: 617-919-2735 F: 617-730-0247 E: Theresa.applegate@childrens.harvard.edu From: Gindhart, Joe (NIH/NIGMS) [E] To: Urdaneta, Angela (NIH/NIGMS) [E] Subject: RE: 2R01GM069808 - 09A1 PI Name: Schwarz, Thomas L. (BCH) Date: Monday, August 25, 2014 9:11:41 AM Attachments: SF424_RR_Guide_ModifiedScopeTemplate.doc Angela, I approve of this request, but they need to complete the attached form, and then have the AOR send it back. Thanks, Joe From: Urdaneta, Angela (NIH/NIGMS) [E] Sent: Friday, August 22, 2014 3:51 PM To: Gindhart, Joe (NIH/NIGMS) [E] Subject: FW: 2R01GM069808 - 09A1 PI Name: Schwarz, Thomas L. (BCH) Dear Joe, Please review and provide programmatic approval if you consider so. Thanks in advance, Angela Urdaneta, MBA | Grants Management Specialist | National Institutes of Health | National Institute of General Medicine Sciences | T: 301-451-2005 | F: 301-480-2554 | Email: urdanetaa@mail.nih.gov From: Applegate, Theresa [mailto:Theresa.Applegate@childrens.harvard.edu] Sent: Friday, August 22, 2014 3:49 PM To: Urdaneta, Angela (NIH/NIGMS) [E];Gindhart, Joe (NIH/NIGMS) [E] Cc: Schwarz, Thomas Subject: 2R01GM069808 - 09A1 PI Name: Schwarz, Thomas L. (BCH) Dear Dr. Gindhardt and Ms. Urdaneta, We received the news that the competing renewal of Dr. Schwarzs Milton and the Transport of Mitochondria project (2R01GM069808 - 09A1) will be awarded, but at a reduced rate of funding. Therefore, we are requesting permission to change the scope of work by removing Specific Aim 1C. Dr. Schwarzs commitment to the project remains unchanged at 1.8 calendar months effort. Attached, please find a copy of Dr. Schwarzs original request along with updated versions of the Project Summary/Abstract and the Specific Aims sections of the application. Should you have any questions or if you need additional information, please do not hesitate to contact me. Thank you for your continued support. Best regards, Theresa Theresa Applegate Boston Childrens Hospital Manager of Sponsored Programs 300 Longwood Avenue Boston, MA 02115 P: 617-919-2735 F: 617-730-0247 E: Theresa.applegate@childrens.harvard.edu From: Urdaneta, Angela (NIH/NIGMS) [E] To: osp@childrens.harvard.edu Cc: thomas.schwarz@childrens.harvard.edu;gindhartjg@mail.nih.gov Subject: Grant Number: 2R01GM069808 - 09A1 PI Name: Schwarz, Thomas L. Date: Tuesday, August 19, 2014 9:13:00 AM Importance: High Dear Authorized Official Representative, We are in receipt of correspondence from the PI requesting to modify the research proposal due to budgetary cuts. To make it official, could your institution please submit the request clearly defining what its now being proposed, and make sure to follow the guidelines in the Grants Policy Statement 8.1.2.5 for the prior approval letter. I would appreciate if the letter is submitted by COB 08/22/14 since we are approaching the end of FY14 and we need to obligate all the funds. Best regards and let me know if you have any questions, Angela Urdaneta, MBA | Grants Management Specialist | National Institutes of Health | National Institute of General Medicine Sciences | T: 301-451-2005 | F: 301-480-2554 | Email: urdanetaa@mail.nih.gov

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
2R01GM069808-09A1
Application #
8759921
Study Section
Synapses, Cytoskeleton and Trafficking Study Section (SYN)
Program Officer
Gindhart, Joseph G
Project Start
Project End
Budget Start
Budget End
Support Year
9
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Children's Hospital Boston
Department
Type
DUNS #
City
Boston
State
MA
Country
United States
Zip Code
02115
Pekkurnaz, Gulcin; Trinidad, Jonathan C; Wang, Xinnan et al. (2014) Glucose regulates mitochondrial motility via Milton modification by O-GlcNAc transferase. Cell 158:54-68
Ashrafi, Ghazaleh; Schlehe, Julia S; LaVoie, Matthew J et al. (2014) Mitophagy of damaged mitochondria occurs locally in distal neuronal axons and requires PINK1 and Parkin. J Cell Biol 206:655-70
Cronin, Michelle A; Schwarz, Thomas L (2012) The CAP-Gly of p150: one domain, two diseases, and a function at the end. Neuron 74:211-3
Wang, Xinnan; Winter, Dominic; Ashrafi, Ghazaleh et al. (2011) PINK1 and Parkin target Miro for phosphorylation and degradation to arrest mitochondrial motility. Cell 147:893-906
Wang, Xinnan; Schwarz, Thomas L (2009) The mechanism of Ca2+ -dependent regulation of kinesin-mediated mitochondrial motility. Cell 136:163-74
Wang, Xinnan; Schwarz, Thomas L (2009) Imaging axonal transport of mitochondria. Methods Enzymol 457:319-33
Goldstein, Ann Y N; Wang, Xinnan; Schwarz, Thomas L (2008) Axonal transport and the delivery of pre-synaptic components. Curr Opin Neurobiol 18:495-503
Glater, Elizabeth E; Megeath, Laura J; Stowers, R Steven et al. (2006) Axonal transport of mitochondria requires milton to recruit kinesin heavy chain and is light chain independent. J Cell Biol 173:545-57