A precise and versatile platform for rapid glycosylation analysis of brain tissue

Vacchini, Mattia; Cipolla, Laura; Gornik, Olga; Lauc, Gordan

doi:10.1039/c9ay02596k

Cited by 5 publications

(2 citation statements)

References 34 publications

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“…Fluorescently labeled N-glycans were separated by HILIC on an Acquity UPLC instrument (Waters, USA) as described previously ( 14 , 63 ) with excitation and emission wavelengths of 250 and 428 nm, respectively. The instrument was under the control of Empower 3 software, build 3471 (Waters).…”

Section: Methodsmentioning

confidence: 99%

Human-specific features and developmental dynamics of the brain N-glycome

Klarić,

Gudelj,

Santpere

et al. 2023

Sci. Adv.

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Comparative “omics” studies have revealed unique aspects of human neurobiology, yet an evolutionary perspective of the brain N-glycome is lacking. We performed multiregional characterization of rat, macaque, chimpanzee, and human brain N-glycomes using chromatography and mass spectrometry and then integrated these data with complementary glycotranscriptomic data. We found that, in primates, the brain N-glycome has diverged more rapidly than the underlying transcriptomic framework, providing a means for rapidly generating additional interspecies diversity. Our data suggest that brain N-glycome evolution in hominids has been characterized by an overall increase in complexity coupled with a shift toward increased usage of α(2-6)–linked N -acetylneuraminic acid. Moreover, interspecies differences in the cell type expression pattern of key glycogenes were identified, including some human-specific differences, which may underpin this evolutionary divergence. Last, by comparing the prenatal and adult human brain N-glycomes, we uncovered region-specific neurodevelopmental pathways that lead to distinct spatial N-glycosylation profiles in the mature brain.

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Section: Methodsmentioning

confidence: 99%

Human-specific features and developmental dynamics of the brain N-glycome

Klarić,

Gudelj,

Santpere

et al. 2023

Sci. Adv.

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“…The cell pellets were thawed before proceeding with a previously described lysate insolution deglycosylation protocol (Vacchini et al, 2020). Briefly, the cell pellets were lysed and glycoproteins extracted using chloroform and methanol.…”

Section: N-glycosylation Analysis Of Harvested Cells Deglycosylation ...mentioning

confidence: 99%

Total cell N-glycosylation is altered during neuronal differentiation of iPSC to NSC and is disturbed by trisomy 21

Cindrić

Vučković

Murray

et al. 2023

Preprint

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Down syndrome (DS) is a genetic condition caused by trisomy 21 (T21) and characterized by a range of neurological symptoms including intellectual disability, early onset of neurodegeneration and dementia, some of which can be attributed to accelerated aging. N-glycosylation is a protein modification that plays a critical role in numerous biological processes and its dysregulation is associated with a wide range of diseases, in some even before the onset of symptoms. N-glycosylation of total plasma proteins, as well as specific plasma proteins, such as immunoglobulin G, has been shown to change in DS, displaying an accelerated aging phenotype consistent with the various symptoms of premature aging that occur in DS. However, little is known about how T21 affects the N-glycosylation of other cellular proteins. To better understand how T21 affects N-glycosylation during neural differentiation, we characterized and compared the total released N-glycans of induced pluripotent stem cells (iPSCs) and their neural stem cell (NSC) derivatives. We analyzed six different isogenic clones all derived from a single individual with mosaic DS and thus all sharing the same genetic background; however, three had a normal disomic karyotype (D21), while the other three had an additional copy of chromosome 21 (T21). We characterized the total cell N-glycosylation profiles using ultra high performance liquid chromatography (UHPLC) and subsequent tandem mass spectrometry analysis to determine proposed glycan structures. Our results revealed both qualitative and quantitative differences in the composition of N-glycomes between iPSCs and NSCs, with NSCs showing a higher amount of complex N-glycans and a lower amount of mannosidic N-glycans when compared to iPSCs. Moreover, we found differences in N-glycosylation patterns between D21 and T21 cells. Notably, T21 cells exhibited a significant increase in the amount of pseudohybrid N-glycans. Trisomy 21 also caused a significant decrease in the abundance of a hybrid monoantennary fucosylated glycan (H6N3F1). Our findings define the released N-glycan profile of total cells for both D21 and T21 iPSCs and NSCs and suggest that the presence of a third copy of chromosome 21 impacts N-glycosylation patterns already in the stem cell state.

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The dynamic brain N-glycome

Lauc

2022

Glycoconj J

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A precise and versatile platform for rapid glycosylation analysis of brain tissue

Abstract: Lysate in-Solution Deglycosylation (LSD) is a novel method for the preparation of N-glycans from tissue. The substrate for LSD is a commonly used lysate that is the starting point for many other applications, including classical proteomics methods.

Cited by 5 publications

References 34 publications

Human-specific features and developmental dynamics of the brain N-glycome

Human-specific features and developmental dynamics of the brain N-glycome

Total cell N-glycosylation is altered during neuronal differentiation of iPSC to NSC and is disturbed by trisomy 21

The dynamic brain N-glycome

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