# Neurobiological significance of Aqp4 stop codon readthrough

> **NIH NIH R00** · UNIVERSITY OF TEXAS DALLAS · 2021 · $249,000

## Abstract

Project Summary/Abstract
Soluble amyloid beta (Aβ) oligomers trigger tau tangle formation, neuronal cell loss, synaptic dysfunction and
cognitive decline seen in Alzheimer's disease (AD). The water channel Aquaporin 4 (Aqp4) is a key component
of the Aβ removal machinery in the brain, as evidenced by ~ 55% reduction in Aβ removal in Aqp4-/- mice.
Specifically, astrocyte endfeet-concentrated Aqp4 is shown to be both required for removing Aβ during sleep
and perturbed in AD, suggesting that restoring Aqp4 to endfeet can improve the outcome of AD. I find that this
endfeet-localized Aqp4 is a stop codon readthrough version of Aqp4. I performed ribosome footprinting (RF),
deep-sequencing of ribosome-protected mRNA fragments, in the mouse brain and detected reads mapping to
the 3' untranslated region of Aqp4, suggesting that ribosomes read past the stop codon and make a C-
terminally extended version of Aqp4 (Aqp4X hereafter). Using an antibody against the readthrough epitope, I
show that Aqp4X is exclusive to the perivascular endfeet, whereas the normal un-extended Aqp4 is confined
elsewhere along the astrocyte membrane. Therefore, the objective of this project is to determine if Aqp4
readthrough enhances Aβ clearance and thus improve AD outcome.
 I propose 3 aims to meet this objective. In aim 1, I will determine if Aqp4X has altered efficacy in eliminating
Aβ compared to Aqp4. I will express either Aqp4X or Aqp4 using viral transduction in the hippocampi of
APP/PS1+/- transgenic mice, and use a novel micro-immunoelectrode technology to measure the rate of Aβ
removal from the interstitial fluid in live mice. Next, on the Aqp4-/- nice that I have acquired and Aqp4No_X
mice that I have generated, I will use ELISA to measure their total brain Aβ levels, with or without viral
rescue. I will also examine Aqp4No_X mice for memory and other behavioral deficits. In aim 2, I will identify the
chemical and genetic regulators of Aqp4 readthrough using drug screening and CRISPRi screening,
respectively. Finally, in aim 3, as an independent investigator, I will determine the AD-related
pathophysiological consequences arising from the loss of endfeet Aqp4. To this end, I will examine Aqp4No_X
mice for possible structural and functional defects in the BBB and neuronal-activity dependent gene regulation
in the hippocampus. I will also cross these mice with APP/PS1 mice and test if Aβ burden and behavioral
deficits escalate when an AD mouse loses endfeet Aqp4. Thus, aim 1 will test the necessity and sufficiency of
the two Aqp4 versions in Aβ clearance, aim 2 will allow future studies on potential therapeutics and biological
regulators, and aim 3 will further elucidate the role Aqp4X plays in AD.

## Key facts

- **NIH application ID:** 10409864
- **Project number:** 4R00AG061231-03
- **Recipient organization:** UNIVERSITY OF TEXAS DALLAS
- **Principal Investigator:** Darshan Sapkota
- **Activity code:** R00 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $249,000
- **Award type:** 4N
- **Project period:** 2021-07-15 → 2024-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10409864

## Citation

> US National Institutes of Health, RePORTER application 10409864, Neurobiological significance of Aqp4 stop codon readthrough (4R00AG061231-03). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10409864. Licensed CC0.

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