Hartnup Disease-Causing SLC6A19 Mutations Lead to B0AT1 Aberrant Trafficking and ACE2 Mis-localisation Implicating the Endoplasmic Reticulum Protein Quality Control

Nesreen F Alkhofash¹¹Bassam R Ali^2*

• ¹United Arab Emirates University, Al-Ain, United Arab Emirates
• ²College of Medicine and Health Sciences, United Arab Emirates University, AlAin, Abu Dhabi, United Arab Emirates

The interaction between angiotensin-converting enzyme 2 (ACE2) and the sodium-dependent Broad neutral Amino acid Transporter 1 (B0AT1) has emerged as a significant focus of research in understanding various physiological and pathological processes. B0AT1, encoded by the SLC6A19 gene, is involved in the neutral amino acid transport across cell membranes and hypothesized to play a role in nutrient absorption. Crucially, several variants in the SLC6A19 gene have been implicated in Hartnup disease, an inborn error of neutral amino acid transport characterized by aminoaciduria and, in some cases, a pellagra-like rash, ataxia, and psychotic behaviour. Conversely, ACE2, an essential gene primarily recognized for its role in the renin-angiotensin system, is responsible for converting angiotensin II (Ang II) into angiotensin 1-7 (Ang 1-7), a peptide that plays crucial roles in vascular homeostasis and inflammation modulation. Furthermore, it has been reported that ACE2 is involved B0AT1 cell surface expression. Recent studies have suggested that ACE2-B0AT1 interactions could influence amino acid transport dynamics, impacting both cardiovascular health and metabolic functions. In addition, the underlying cellular mechanisms of Hartnup disease-causing mutations remain unknown. Therefore, in this study, we evaluated the subcellular trafficking and targeting of 18 Hartnup Disease-causing B0AT1 (SLC6A19) variants. Our results indicated that 9 of these variants (R57C, G93R, R95P, R178Q, L242P, G284R, S303L, D517G, P579L) resulted in endoplasmic reticulum (ER) retention of the mutant protein, preventing or retarding its trafficking to the plasma membrane. These variants are distributed across various B0AT1 structural domains. We also investigated their potential effects on ACE2 subcellular trafficking and targeting using experimental approaches involving subcellular localization and biochemical analysis. Our data show that the 9 ER-retained B0AT1 disease-causing missense variants affected ACE2 localization to varying degrees. In particular, R178Q and S303L variants showed significant impact on ACE2 intracellular trafficking and subcellular targeting to the plasma membrane. Establishing the pathogenesis mechanisms underlying Hartnup disease as well as improving our understanding of the molecular mechanisms of ACE2 and B0AT1 protein interactions, could open new avenues for understanding disease pathogenesis. It may also offer opportunities for therapeutic intervention and biomarker development in associated conditions, which requires further research.