# Role of FK506-Binding Protein 3 (FKBP3) as a Novel Regulator of Skeletal Muscle Protein Synthesis

> **NIH NIH R21** · INDIANA UNIVERSITY INDIANAPOLIS · 2022 · $209,220

## Abstract

Ca2+/calmodulin-dependent protein kinase kinase 1 (CaMKK1) has been implicated in the regulation of
skeletal muscle protein synthesis via a mechanistic target of rapamycin complex 1 (mTORC1)-dependent
mechanisms. However, the downstream signaling proteins/substrates that connect CaMKK1 to mTORC1 to
initiate this beneficial effect on muscle protein homeostasis are currently unknown. The long-term goal of this
application is to develop novel drug therapies for muscle atrophy that target constituents of the signaling pathway
utilized by CaMKK1 to stimulate muscle protein synthesis. The overall hypothesis is that the phosphorylation
status of FKBP3 controls its isomerase activity and subcellular localization; and that these functional and spatial
changes facilitate the ability of FKBP3 to regulate mTORC1 signaling and protein synthesis in skeletal muscle.
The rationale is based in part on an unbiased, ATP-analog sensitive kinase and quantitative phospho-proteomics
approach showing that CaMKK1 phosphorylates FKBP3 on T122, a residue located in its peptidyl-prolyl
isomerase domain. To test the overall hypothesis, the following two specific aims were proposed: 1) Determine
if FKBP3 regulates mTORC1 signaling and protein synthesis in muscle, and 2) Determine if FKBP3 isomerase
activity or localization is regulated by T122 phospho-status. The first aim will use in vivo muscle gene
transfer/electroporation to express wild-type and T122 phospho-site mutant proteins in the muscle of male &
female mice. The effects of FKBP3 phospho-status on changes in muscle mass/cross-sectional area, mTORC1
signaling proteins, and protein synthesis, will be assessed. The second aim will use purified wild-type CaMKK1
and FKBP3 proteins, cell-free phosphorylation assays, and mass spectrometry to identity all possible CaMKK1-
stimulated phospho-sites on FKBP3. Next, FKBP3 T122 phospho-mutant proteins will be generated, and
peptidyl-prolyl isomerase activity examined in the presence or absence of rapamycin. We will also use in vivo
electroporation to express active CaMKK1 in mouse muscle and then examine FKBP3 isomerase. Last, we will
express fluorescently labeled wild-type and T122 phospho-mutant FKBP3 in mouse muscle, and then assess
FKBP3 subcellular localization by fluorescence microscopy. The proposed research is innovative because it will
determine whether FKBP3 is part of a novel Ca2+/CaMKK1-dependent mechanism that regulates mTORC1
signaling and protein synthesis in skeletal muscle. The proposed research is significant because it will define a
part of the cellular mechanism that links intracellular Ca2+ to mTORC1 signaling in muscle. This is a key first step
towards the generation of new therapies for muscle atrophy that would target this signaling pathway.

## Key facts

- **NIH application ID:** 10517563
- **Project number:** 1R21AR081593-01
- **Recipient organization:** INDIANA UNIVERSITY INDIANAPOLIS
- **Principal Investigator:** Carol Ann Witczak
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $209,220
- **Award type:** 1
- **Project period:** 2022-07-28 → 2024-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10517563

## Citation

> US National Institutes of Health, RePORTER application 10517563, Role of FK506-Binding Protein 3 (FKBP3) as a Novel Regulator of Skeletal Muscle Protein Synthesis (1R21AR081593-01). Retrieved via AI Analytics 2026-06-11 from https://api.ai-analytics.org/grant/nih/10517563. Licensed CC0.

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