The research team led by Director C Justin LEE of the Center for Cognition and Sociality within the Institute for Basic Science (IBS), along with the team led by Chief Investigator KIM Ho-Min of the Center for Biomolecular and Cellular Structure, has elucidated the operational mechanism of a key ion channel crucial for maintaining the shape and function of the Golgi apparatus, an organelle responsible for the modification and transport of proteins within cells. The study also revealed the relationship between damage to this ion channel in brain cells and cognitive impairment, presenting new therapeutic targets for brain diseases.

The Golgi apparatus acts like a post office within cells, receiving lipids and proteins synthesized by the endoplasmic reticulum, processing and modifying them, and then transporting them to other cellular organelles or outside the cell. For the Golgi apparatus to maintain its shape and function, it is important for the internal environment to be maintained at a mildly acidic pH of 6.0-6.7 by ion channels. Malfunctions in these ion channels are known to cause structural changes in the Golgi apparatus, which is a condition often found in Alzheimer’s disease patients. However, the relationship between Golgi structure and cognitive impairment was not clear due to a lack of understanding of these ion channels.

In a quest to identify major ion channels in the brain cell Golgi apparatus, Director C Justin LEE's team explored various uncharacterized membrane proteins. In 2023, they identified that the membrane protein TMEM87A, which is highly expressed in hippocampal astrocytes and neurons, regulates the acidity inside the Golgi apparatus of brain cells by acting as a cation channel. They named this membrane protein "Golgi-pH-regulating cation channel" or "GolpHCat" to emphasize its physiological role.

In collaboration with Chief Investigator KIM Ho Min's team, they used IBS's cryo-electron microscopy (Cryo-EM) to determine the three-dimensional molecular structure of GolpHCat at an ultra-high resolution of 3.1 Ångströms (1/100 million cm). They also conducted electrophysiological experiments and molecular dynamics analyses to reveal the ion movement pathway. It was established that GolpHCat is a voltage-dependent channel that opens in response to voltage changes across the cell membrane.

GolpHCat has a structure comprising a lumen domain connected to the cell membrane on the external side and a transmembrane domain with seven helices penetrating the membrane, with a cavity in the center of the transmembrane domain. This cavity is physically blocked by fatty acid chains of phosphatidylethanolamine originating from the Golgi membrane, preventing ion passage. However, when voltage is applied, the third helix of the transmembrane domain acts as a voltage sensor, activating and opening the channel. The negatively charged residues inside the funnel-shaped lumen then attract positively charged sodium (Na), potassium (K), and cesium (Cs) ions into the channel. Thus, GolpHCat serves as a pathway for cations, contributing to the maintenance of Golgi membrane voltage and internal acidity in conjunction with anion channels.

To confirm the biological effect that arises due to GolpHCat damage, the research team observed brain cells from GolpHCat-deficient mice. These mice exhibited abnormal structural changes such as the Golgi apparatus fragmenting or swelling. These structural changes disrupted one of the Golgi's main functions, protein glycosylation, leading to impaired learning and memory in the mice. These findings highlight the importance of normal GolpHCat function for cognitive abilities and suggest the potential for targeting GolpHCat in treating cognitive disorders.

Chief Investigator KIM Ho Min remarked, "This research is a result of a convergence study incorporating various advanced bio research technologies such as neurobiology, structural biology, molecular dynamics, and glycomics, representing a successful case of collaboration that broke down academic barriers."

Director C Justin LEE stated, "We have elucidated how structural and functional changes in the Golgi apparatus are involved in memory. Understanding the molecular mechanisms of the Golgi apparatus could lead to new treatments for cognitive impairments found in various neurodegenerative brain diseases."

This research, which discovered a new Golgi ion channel and identified its function, structure, and physiological importance, represents a collaborative success within the IBS Life Science Institute. The study was also conducted in collaboration with Professor CHOI Sun Young from Ewha Womans University's College of Pharmacy (supported by the Ministry of Science and ICT and the National Research Foundation of Korea), and Professor AN Hyun Joo from Chungnam National University's Department of Analytical Science and Technology.

The results of the study were published online in the journal "Nature Communications" (IF=14.7) on July 11.

Figure 1. Schematic of Molecular Structure Study of GolpHCat
Using cryo-electron microscopy, the molecular structure of human GolpHCat was elucidated at high resolution, revealing that GolpHCat is an ion channel with a unique structure composed of two domains in a monomeric form. Additionally, the complex structure with a pharmacological inhibitor (gluconate) and the structure of mutants that interfere with phospholipid binding were identified. Based on these molecular structures, electrophysiological experiments and molecular dynamics modeling experiments were conducted to elucidate the ion transport pathway within the monomer and the mechanism of voltage-gated channel opening and closing. Specifically, it was found that negatively charged residues inside the funnel-shaped lumen of GolpHCat serve as entry points that attract cations. It was confirmed that the pharmacological inhibitor blocks an important ion transport pathway inside GolpHCat (top right). Furthermore, molecular dynamics experiments identified the phospholipid binding pathway and various binding forms of phospholipids, proposing the mechanism of voltage sensing and voltage-gated channel opening and closing in GolpHCat (bottom right).

Figure 2. GolpHCat is Important for Maintaining Normal Golgi Structure
As a cation channel that regulates Golgi pH, GolpHCat is crucial for maintaining the normal structure of the Golgi apparatus in astrocytes and neurons in the hippocampus. The Golgi structure in astrocytes and neurons in the hippocampus was observed using transmission electron microscopy. In GolpHCat knockout (KO) mice, the maximum length of Golgi cisternae in astrocytes and neurons was reduced, and the width of the Golgi was increased compared to the control group (WT), showing fragmented or swollen structures. GolpHCat is involved in maintaining the normal structure of the Golgi apparatus in astrocytes and neurons in the hippocampus.

Figure 3. Memory Impairment in GolpHCat Knockout Mice
Memory dependent on the hippocampus was found to be reduced in GolpHCat knockout (KO) mice expressing defective GolpHCat in astrocytes and neurons. (Left) The novel place recognition test, which evaluates spatial memory in mice, involves observing the exploration of two objects placed in a box, then moving one object to a new location after a certain period to see if the mice remember the original location (space) of the objects. GolpHCat KO mice showed increased exploration time of the familiar object and decreased exploration time of the object in the new location compared to the control group (WT), indicating impaired recognition and spatial memory. (Right) The contextual fear conditioning test, which evaluates contextual memory in mice, involves giving a mild electric shock through the floor of a test box and then observing if the mice show a fear response (freezing) when placed back in the test box after a significant period. GolpHCat KO mice showed reduced fear response time compared to the control group (WT), indicating impaired contextual memory.