Endolysosomal Cation Channels of the Transient Receptor Potential Superfamily

Table of Contents

1 List of original publications selected for this „Habilitationsschrift“

2 Introduction

3 Analysis of the TRPML3 varitint-waddler mouse mutants

4 Development of small molecule agonists for TRPML channels

5 Functional investigation of MLIV causing TRPML1 mutants

6 Novel TRPML channel interaction partners

7 Functional analysis of the TPC2-/- knockout mouse

8 Novel TPC channel interaction partners

9 Summary

10 References

11 Curriculum vitae

12 Complete list of original publications

13 Personal contributions to the selected original publications

14 Acknowledgement

Objectives and Research Focus

This thesis investigates the physiological roles and functional mechanisms of endolysosomal cation channels from the TRP superfamily, specifically focusing on TRPML1, TRPML3, and TPC2. The research aims to elucidate how these channels contribute to intracellular trafficking, ion homeostasis, and the pathophysiology of hereditary disorders such as mucolipidosis type IV, ultimately exploring their potential as pharmacological targets.

• Mechanistic analysis of gain-of-function mutations in TRPML3 and their impact on hair cell survival.
• High-throughput identification and development of small molecule agonists for TRPML channel activation.
• Restoration of channel function in TRPML1 mutants associated with mucolipidosis type IV.
• Identification of novel protein-protein interaction partners, including TMEM163 and syntaxins, to map channel-mediated cellular processes.
• Phenotypic characterization of TPC2-deficient mouse models regarding hepatic cholesterol metabolism and intracellular trafficking.

Excerpt from the Book

Summary of Chapters

1 List of original publications selected for this „Habilitationsschrift“: Provides an overview of the key peer-reviewed scientific articles central to this habilitation.

2 Introduction: Establishes the current knowledge regarding TRP and TPC channels in endolysosomal systems and their association with human diseases.

3 Analysis of the TRPML3 varitint-waddler mouse mutants: Examines the molecular mechanism by which A419P mutations induce constitutive TRPML3 activity and subsequent cellular degeneration.

4 Development of small molecule agonists for TRPML channels: Details the high-throughput screening process to identify novel pharmacological activators for TRPML channels.

5 Functional investigation of MLIV causing TRPML1 mutants: Discusses strategies to restore channel function in patient-derived fibroblasts using small molecule ligands.

6 Novel TRPML channel interaction partners: Identifies TMEM163 as an interacting protein that regulates cellular zinc homeostasis.

7 Functional analysis of the TPC2-/- knockout mouse: Analyzes the impact of TPC2 loss on macromolecular trafficking and liver-specific cholesterol pathology.

8 Novel TPC channel interaction partners: Explores the role of SNARE proteins, specifically STX7, in TPC2-mediated vesicle fusion processes.

9 Summary: Synthesizes the primary findings across all presented publications regarding channel physiology and disease modeling.

10 References: Lists the scientific literature cited throughout the work.

11 Curriculum vitae: Outlines the academic and professional career of the author.

12 Complete list of original publications: A comprehensive bibliography of the author's peer-reviewed scientific articles, reviews, and book chapters.

13 Personal contributions to the selected original publications: Clarifies the specific experimental and intellectual contributions of the author to each cited study.

14 Acknowledgement: Expresses gratitude to mentors, colleagues, and family members for their support.

Key Terms

TRPML, TPC, Ion Channels, Endolysosomal System, Mucolipidosis Type IV, Varitint-Waddler, Small Molecule Agonists, Patch-Clamp, Intracellular Trafficking, Zinc Homeostasis, Cholesterol Metabolism, SNARE Proteins, Constitutive Activity, Mutation Analysis, Hearing Loss

Frequently Asked Questions

What is the primary focus of this research?

This work explores the physiological functions, structural mechanisms, and potential therapeutic targeting of cation channels within the endolysosomal system, specifically members of the TRPML and TPC families.

What are the central thematic fields addressed in this work?

The core themes include membrane trafficking, ion homeostasis, the molecular pathology of lysosomal storage disorders, and the identification of pharmacological tools to modulate channel activity.

What is the primary scientific goal or research question?

The research aims to understand how mutations in endolysosomal channels lead to disease phenotypes and whether targeting these channels with small molecules or identifying their protein interaction partners can help restore cellular function.

Which scientific methods are utilized in this thesis?

The study employs a broad range of techniques, including patch-clamp electrophysiology, high-throughput small molecule screening, confocal microscopy, calcium imaging, and the development of knockout mouse models.

What does the main body of the work cover?

It provides a deep dive into the characterization of TRPML3 mutations, the development of small molecule agonists for TRPML channels, the restoration of TRPML1 function in patient cells, and the interaction of these channels with the endolysosomal machinery.

Which keywords best characterize this work?

Essential keywords include TRPML, TPC, endolysosomal system, ion channels, mucolipidosis type IV, intracellular trafficking, and small molecule pharmacology.

What role does TRPML3 play in the hearing-related phenotypes discussed?

The study demonstrates that the A419P mutation in TRPML3 renders the channel constitutively active, causing a massive calcium and sodium overload that results in hair cell death and subsequent deafness.

What is the significance of the findings regarding TPC2 in liver health?

The analysis of TPC2 knockout mice reveals a crucial role for the channel in endolysosomal trafficking of cholesterol; its loss renders the mice highly susceptible to cholesterol accumulation, fatty liver disease, and gallstone formation.