Researchers identify a new gene involved in the rare lysosomal storage disorder

Researchers identify a new gene involved in the rare lysosomal storage disorder

A rare disease called mucolipidosis type II causes the heart and abdomen to swell and the bones to deform.

Mucolipidosis type II is a lysosomal storage disorder and causes edema of internal organs and skeletal dysplasia. Children diagnosed with the genetic disease often die before they reach the age of seven. Now researchers at the University of Michigan have identified a new gene involved in the disease: TMEM251, which is necessary for the correct function of lysosomes.

Lysosomes are organelles in all cells in the body - except red blood cells - that are responsible for absorbing and recycling the waste your cells produce. If the lysosome cannot function properly, it cannot recycle this waste and instead simply stores it in the organelle.

The team, led by Ming Li, assistant professor of molecular, cellular and developmental biology, discovered that when TMEM251 is defective, it is unable to encode the pathway for the enzymes required for the correct function of lysosomes to move within the lysosome. The study was published in Nature Communications.

Lysosomes contain around 50 to 60 enzymes that digest worn-out cell parts and waste from outside the cell. The lysosome also recycles these wastes – proteins, nucleic acids, carbohydrates and lipids – back into usable material. However, in order for these enzymes to move within the lysosome, they require a signal called the mannose-6-phosphate biosynthetic pathway, or M6P.

It's like a postage stamp. The enzymes must have this signal in order to enter the lysosome. If they don't have M6P, they can't get into the lysosome. “As a result, there are still lysosomes, but none of them would be functional because they lack these enzymes.”

Ming Li, assistant professor of molecular, cellular and developmental biology

Li's laboratory studies the lysosome and, in particular, the composition of the lysosome's membrane proteins. The lysosome has the ability to regulate its own membrane proteins by triggering the breakdown of these proteins through a process called ubiquitination. This process allows proteins to migrate from the lysosome membrane into the organelle and be broken down there. The researchers also wanted to understand which genes are responsible for lysosome function and what happens when these genes are defective.

To do this, the team used a CRISPR knockout screen that individually switched off each gene in the human genome at the cellular level. The researchers were then able to examine what happens in the lysosome in response to the deletion of each gene. Specifically, the researchers looked for genes that could be responsible for the breakdown of the lysosome.

The experiment yielded TMEM251.

"Then the question became: Why is this gene so important for human health? And why is it so important for lysosomal function?" Li said.

The group discovered that the TMEM251 gene encodes an enzyme that activates M6P, a pathway required by most of the 50 to 60 digestive enzymes in lysosomes. In a literature search, the researchers also found a 2021 paper that described mucolipidosis type II-like symptoms in humans that are due to a defective TMEM251 gene.

“Our discovery answered the molecular mechanism of this new human disease,” Li said.

The protein encoded by the TMEM251 gene is required to activate another enzyme called GNPT, which catalyzes the M6P pathway. The researchers also showed that TMEM251 is localized to the Golgi apparatus, a structure that forms lysosomes. That the two enzymes are located at the Golgi fits with the idea that the proteins must work together to add M6P to lysosomal enzymes, Li said. The researchers named TMEM251 the GNPT cleavage and activity factor (GCAF).

The researchers then checked what would happen if they turned off the TMEM251 gene in zebrafish. By comparing the wild-type zebrafish with the zebrafish whose TMEM251 gene had been knocked out, the researchers were able to detect defects in the zebrafish's abdomen, skeletal and cartilage development, and heart.

Co-author Xi Yang said the team also proposes a therapeutic strategy to combat the disease in humans. The therapy, which is in a very early stage, is based on a so-called “enzyme replacement therapy”. The researchers showed that when they delivered the enzyme containing the M6P modification to TMEM251-deficient cells, this enzyme was able to enter the cell through a process called endocytosis and be delivered to a malfunctioning lysosome.

"We know that the pathogenesis of this disease comes from not having a functional lysosome," said Yang, a research specialist in Li's lab. "This knockout cell can actually use these endocytosed functional enzymes to rebuild its lysosome and make it functional again. You can correct the deficiency, at least at the cellular level."

The team recently received a grant from the National Institutes of Health to further study the TMEM251 gene, specifically how the TMEM251 enzyme interacts with the GNPT enzyme to facilitate the formation of M6P. The team also wants to describe what TMEM251 looks like at a structural level.

Co-authors of the paper include Weichao Zhang, Linchen Yu, Bokai Zhang, Jianchao Zhang, Varsha Venkatarangan, Liang Chen, Sarah Bui and Yanzhuang Wang from UM MCDB. MCDB professor Cunming Duan and postdoctoral fellow Yingxiang Li helped with the zebrafish work. Woo Yung Cho joined the team from the BRCF Microscopy Core at UM Medical School. Bala Bharathi Burugula and Jacob Kitzman from the Department of Human Genetics at UM Medical School also contributed.

Source:

University of Michigan

Reference:

Zhang, W., et al. (2022) GCAF(TMEM251) regulates lysosome biogenesis through activation of the mannose-6-phosphate pathway. Nature communication. doi.org/10.1038/s41467-022-33025-1.

.