# Administrative Supplement for Unraveling the Mechanisms of Neurodegeneration in TBCK Encephaloneuronopathy

> **NIH NIH R01** · CHILDREN'S HOSP OF PHILADELPHIA · 2024 · $107,662

## Abstract

Abstract
The disruption of the homeostasis in neurons often results in degeneration, an irreversible mechanism that still
is not well understood. Neurodegeneration is often accompanied by lysosomal dysfunction, which is
characterized by increased luminal pH levels, alterations to the proteolytic capacity, and disruption of the activity
of lysosomal transmembrane proteins. Fibroblasts and induced neurons from the most severe variant (p.R126X)
of a rare pediatric-onset neurodevelopmental disease called TBCK syndrome show a strong phenotype
associated with lysosomal dysfunction; however, the mechanism by which the deficiency of the TBC1-containing
kinase (TBCK) protein alters the effective clearance of senescent or damage proteins and organelles remains
unclear. The p.R126X, also known as the “Boricua” mutation because it affects children from Puerto Rican
ancestry, is a frameshift variant that causes the truncation of the TBCK protein. Proteomic studies performed in
our lab showed that TBCK interacts with the c-Jun N-terminal kinase-interacting protein 4 (JIP4), and the
deficiency of TBCK disrupts the protein levels of JIP4, which also correlate with more stationary lysosomes in
axons from TBCK-deficient induced neurons (TBCK-deficient iNeu). Therefore, I hypothesize that TBCK
deficiency impairs JIP4 stability leading to the disruption of the positioning, motility, and maturation of lysosomes.
The first aim of this project will test the maturation and motility of lysosomes and the role effect of JIP4 disruption
in lysosomal function. To test this, I will define the maturation state of lysosomes of TBCK-deficient iNeu using
a pH-sensitive-LAMP1 reporter and confocal microscopy. Also, I will perform time-lapse live imaging analysis to
assess the motility of lysosome in axons from TBCK-deficient iNeu, and I will disrupt the levels of JIP4 in control
iNeu to observe whether lysosomes reflect the same phenotype as TBCK-deficient iNeu. The second aim will
determine whether introducing a plasmid containing the TBCK sequence into TBCK-deficient iNeu restitutes
JIP4 protein levels and restores the functionality of lysosomes. Also, I will use HEK cells to evaluate which
domain of the TBCK protein, pseudo-kinase, TBC, and rhodanese, interacts with JIP4. This aim will reveal the
role of TBCK in the modulation of JIP4 protein levels and the domain involved on this novel interaction. Overall,
this project will define the relevance of the novel interaction between TBCK and JIP4 and how the disruption of
the levels of JIP4 is implicated in the maturation and motility of lysosomes.

## Key facts

- **NIH application ID:** 10987882
- **Project number:** 3R01NS132795-02S1
- **Recipient organization:** CHILDREN'S HOSP OF PHILADELPHIA
- **Principal Investigator:** XILMA R ORTIZ-GONZALEZ
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $107,662
- **Award type:** 3
- **Project period:** 2023-09-18 → 2028-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10987882

## Citation

> US National Institutes of Health, RePORTER application 10987882, Administrative Supplement for Unraveling the Mechanisms of Neurodegeneration in TBCK Encephaloneuronopathy (3R01NS132795-02S1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10987882. Licensed CC0.

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