Journal Title
Journal ISSN
Volume Title

The congenital disorders of glycosylation (CDG) are a group of diseases with inborn errors in metabolism that result from the improper glycosylation of necessary biological molecules. Though they are a collection of rare diseases, unique CDGs now number in the hundreds. 1 In 2012, N-glycanase deficiency became the world's first known congenital disorder of deglycosylation (CDDG). The clinical presentations of these individuals possess variable expressivity ranging from all patients having global developmental delay, movement disorder and hypotonia, to patients with a mosaic phenotype with the affliction of other neurological (e.g. abnormal brain imaging, electroencephalogram abnormalities, acquired microcephaly, diminished reflexes, seizures) and non-neurological (e.g. hypo- or alacrima, elevated liver transaminases, and hepatocellular storage and vacuolization dysfunction) symptoms. Premature death has also been recorded for several patients with NGLY1 deficiency. 2 As one would assume with the novelty of this disease, molecular mechanisms regarding how the loss of NGLY1 glycanase leads to the widely detrimental effects on human neurological development are mostly unknown. We chose to embark on this journey to elucidate the pathogenesis of this disease, but along the way also asked very pertinent questions revolving around the importance of the NGLY1 protein in other cell types. As an inherited disease, NGLY1 deficiency is a consequence of the mutant NGLY1 gene that has errors in translation within cells from the very beginning of life at the zygote, blastocyst, and other embryogenic stages. Evidenced by the developmental abnormalities observed in NGLY1-deficient patients, the normal function of NGLY1 plays a role in ensuring the normal development of human cells. Given that cancer is characterized as the cell's inability to regulate its proliferation and differentiation capacity, we also hypothesized that the dysregulation of NGLY1 could be involved also in the oncogenic process and cancerous development. Using melanoma as a proof-of-principle cancer type in our studies, the significance of abnormally expressed NGLY1 in human cancer cells has been revealed. In addition, we concluded that NGLY1 represents a novel and promising target that can be suppressed for anti-melanoma effects. Meanwhile, to elucidate the cellular and molecular mechanisms underlying the neurodevelopmental defects due to NGLY1 deficiency, we thought that a human cell-based platform that can closely model the early stage of cerebral development in humans would be necessary to properly and effectively investigate the role of NGLY1 in human brain development. The recent discoveries regarding human cerebral organoids and their ability to self-organize and give rise to cell layers similar to the structure of the human fetal brain made our intended research in the NGLY1-deficient development of the human cerebrum timely and feasible. 3 Organoids established using human induced pluripotent stem cells (hiPSCs) that are reprogrammed from somatic cells harvested from patients that carry pathogenic mutations provide even closer personalized disease modeling for pathogenesis that occurs in the human body. With the goal of building cerebral organoids using hiPSCs reprogrammed from noninvasively collected somatic cells that is suitable for studying neurodevelopment, we decided to test the production of cerebral organoids using hiPSCs reprogrammed from urine samplederived urinary epithelial cells (UECs). Although UEC-derived hiPSCs required specific protocol optimization to properly form cerebral organoids, the cellular and transcriptomic features of these organoids were comparable to those of cerebral organoids developed from embryonic stem cells. Our data offered direct evidence for the suitability, advantage, and possible limitations of using human urine sample-derived COs to model and study neurodevelopment. To tackle the question regarding how NGLY1 deficiency perturbs human brain development, we leverage a platform composed of NGLY1-functional and -deficient cerebral organoids to test our hypothesis that NGLY1 deficiency biases cell fates through compromising stress-adaptation capacity and secretory factor signaling in human cells undergoing neurogenesis. Our studies revealed that, compared with NGLY1-functional cerebral organoids, the NGLY1-deficient ones showed the premature differentiation of neural stem cells (NSCs), higher vulnerability to multiple stressors, and the reduction of secretory factors that are linked to neuroprotective and neurodevelopmental function. We concluded that NGLY1 may play critical roles in regulating stress responses and maintaining NSCs in cerebral development. NGLY1 deficiency-associated neurological abnormalities, including microcephaly, are likely due to the disruption of the NSC signaling modulated by NGLY1.