EXPERIMENT_TITLE: GSM5839926: Hi-C.alpha3; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: Human islets were purchased from Prodo Labs (Cat# HIR-001). After receiving, the islets were well dispersed and washed with complete PIM(S) media (Prodo Labs, Cat# PIM-S001GMP) by centrifuging at 180g for 2 minutes.  The supernatant was removed with about 2-3ml of media left. The islet pellets were then gently resuspended in complete PIM(S) media, and transferred to a 6-well non-tissue culture treated plate. After recovering in a cell incubator for overnight, the islets were digested to single cells. Briefly, the islets were pelleted and washed with HBSS without Ca++/Mg++ (Sigma, Cat# H6648-1L), and then digested with Accutase (Life Technologies, Cat# A1110501) in a 6-well non-treated plate at 37ºC for about 5 minutes. The digested cells were filtered through a 40?m cell strainer to obtain single cells, and the cell number and viability were observed after washing twice with HBSS. To prepare for low input Hi-C, the single cells were firstly fixed with BD cytofix/ cytoperm buffer (BD, Cat# 51-2090KZ) for 20 minutes and washed with BD perm/ Wash buffer before stained with desired antibodies for alpha cells (BD Biosciences, BV421 mouse anti-glucagon, Cat# 565891) and beta cells (BD Biosciences, Alexa Flour 647 Mouse Anti-Insulin, Cat# 565689). The cells were sorted out using BD FACS ARIA II. We followed our previous easy Hi-C protocol to generate the eHi-C libraries. In order to make it compatible to low input sorted cells, we made a few changes. Briefly, about 20-30k sorted alpha or beta cells were applied for each library. Nuclei were extracted and digested with HindIII before proximity ligation. The chromatin ligates were then digested DpnII before constructing self-circles. The circuited DNAs were then linearized with a second digestion of HindIII. All the recovered DNAs were used to generate libraries. We followed the standard protocol to generate the libraries with modified Truseq adapters containing a regular sample index (i7) and a random fragment index (i5: NNNNNN).
STUDY_DESCRIPTOR_STUDY_ABSTRACT: The functional heterogeneity of ß-cells is important for metabolism and diseases, but the nature and mechanism of such variation at molecular level remain elusive. Here we explore this question by comparing both single cell RNA-seq and single cell ATAC-seq maps of healthy and type II diabetic (T2D) human islets. We dissect the T2D-associated single cell trajectory and identified signature genes and enhancers in ß-cells. We also map the 3D genome of both a- and ß-cells with low-input eHi-C approach to unravel the cell type-specific gene regulatory circuits. Strikingly, more than one third of the T2D signature genes show significant intra-donor heterogeneity at single cell level; these genes are functionally distinct from other signature genes that are largely invariant among cells from the same individual but differentially expressed between donors, suggesting different roles in T2D pathogenesis. Importantly, we identified consistent intra-donor variations at both transcriptomic and epigenomic levels, which strongly support the heterogenous ß-cell states and transcription programs. Finally, we construct the disease-signature regulatory networks and pinpointed HNF1A as one of the top transcription factors governing T2D-associated ß-cell heterogeneity. Taken together, we provide the first multi-omic characterization of ß-cell heterogeneity and reveals its connection to T2D. Overall design: 3D genome structure mapping in sorted alpha and beta cells from human islets using easy Hi-C method
SAMPLE_TITLE: Hi-C.alpha3
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "human islet sorted alpha cells"}, {"TAG": "tag", "VALUE": "no treat"}, {"TAG": "tag", "VALUE": "Hi-C"}, {"TAG": "geo_loc_name", "VALUE": "missing"}, {"TAG": "collection_date", "VALUE": "missing"}]
Pool_Member_@attributes_sample_title: Hi-C.alpha3
RUN_SET_RUN_Pool_Member_@attributes_sample_title: Hi-C.alpha3
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