Centromere proteomic analysis of Down syndrome-derived amniocytes

doi:10.3969/j.issn.1674-8115.2026.01.002

Abstract

Abstract:

Objective ·To explore differentially expressed proteins in the centromere proteome of amniocytes from fetuses with Down syndrome (DS). Methods ·Amniocytes from DS and normal fetuses were used as models. A lentiviral vector encoding the centromere protein A-enhanced green fluorescent protein-ascorbate peroxidase 2 (CENPA-EGFP-APEX2) fusion protein was constructed and transfected into amniocytes to target and label centromere-proximal proteins. The expression and location of the target protein were determined by immunofluorescence co-localization, and protein labeling was verified by biotinylation treatment and Western blotting. Quantitative proteomics analysis using 4D label-free technology was used on three DS samples and three normal samples. Functional annotation of the labeled proteins was performed using the Gene Ontology (GO), Clusters of Orthologous Groups/Eukaryotic Orthologous Groups (COG/KOG), and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. Differentially expressed proteins (DEPs) between DS and normal amniocytes were screened with thresholds of P<0.05 and |log₂FC|>0.585, and a protein-protein interaction (PPI) network was constructed using the STRING database to identify core hub proteins. Results ·A centromere-proximal protein labeling model using DS amniocytes was successfully established, and immunofluorescence co-localization and Western blotting confirmed that CENPA-EGFP-APEX2 was precisely anchored at the centromere and labeled proximal proteins. Proteomic analysis identified 999 high-confidence proteins, and bioinformatics analysis showed that these proteins were enriched in redox-related and cancer-related pathways. Differential expression analysis revealed significant upregulation of MX dynamin-like GTPase 1 (MX1) and signal transducer and activator of transcription 1 (STAT1), as well as downregulation of actin β (ACTB). The PPI network further identified small ubiquitin-like modifier 2 (SUMO2), heterogeneous nuclear ribonucleoprotein A/B (HNRNPAB), and mitogen-activated protein kinase (MAPK) as key hub proteins. Conclusion ·There are various differentially expressed proteins in the centromere proteome of DS fetal amniocytes, with abnormal activation of MX1 and STAT1, downregulation of ACTB, and interactions among these differential proteins centered on SUMO2, MAPK, and HNRNPAB.

Key words: Down syndrome (DS), amniocyte, centromere proteome, APEX2 proximity labeling

CLC Number:

R394.3

Zhou Peng, Liu Yanna, Yan Jingbin. Centromere proteomic analysis of Down syndrome-derived amniocytes[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2026, 46(1): 15-24.

Figures/Tables 4

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References 44

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Case	Maternal age/year	Gestation age/week	Fetal sex	Indication for testing	Chromosomal karyotype
1	40	16⁺³	Male	High-risk NIPT result	47, XY, +21
2	30	16⁺⁵	Male	High-risk NIPT result	47, XY, +21
3	44	18⁺²	Female	High-risk NIPT result	47, XX, +21
4	35	18⁺²	Male	High-risk NIPT result	47, XY, +21
5	41	19⁺³	Female	High-risk NIPT result	47, XX, +21
6	31	24⁺⁴	Female	High-risk NIPT result	47, XX, +21
7	32	13	Female	High-risk NIPT result	47, XX, +21
8	31	18⁺³	Male	High-risk NIPT result	47, XY, +21
9	25	16⁺¹	Male	High-risk NIPT result	47, XY, +21
10	38	18	Female	Advanced maternal age	47, XX, +21
11	46	24⁺⁵	Male	Positive screening for trisomy 21	46, XY
12	34	19⁺⁴	Male	Positive screening for trisomy 21	46, XY
13	33	21⁺⁶	Male	Positive screening for trisomy 21	46, XY
14	27	16⁺⁵	Male	High-risk serum markers	46, XY
15	30	17⁺⁵	Female	High-risk serum markers	46, XX
16	42	23⁺³	Male	Advanced maternal age	46, XY

Case	Maternal age/year	Gestation age/week	Fetal sex	Indication for testing	Chromosomal karyotype
1	40	16⁺³	Male	High-risk NIPT result	47, XY, +21
2	30	16⁺⁵	Male	High-risk NIPT result	47, XY, +21
3	44	18⁺²	Female	High-risk NIPT result	47, XX, +21
4	35	18⁺²	Male	High-risk NIPT result	47, XY, +21
5	41	19⁺³	Female	High-risk NIPT result	47, XX, +21
6	31	24⁺⁴	Female	High-risk NIPT result	47, XX, +21
7	32	13	Female	High-risk NIPT result	47, XX, +21
8	31	18⁺³	Male	High-risk NIPT result	47, XY, +21
9	25	16⁺¹	Male	High-risk NIPT result	47, XY, +21
10	38	18	Female	Advanced maternal age	47, XX, +21
11	46	24⁺⁵	Male	Positive screening for trisomy 21	46, XY
12	34	19⁺⁴	Male	Positive screening for trisomy 21	46, XY
13	33	21⁺⁶	Male	Positive screening for trisomy 21	46, XY
14	27	16⁺⁵	Male	High-risk serum markers	46, XY
15	30	17⁺⁵	Female	High-risk serum markers	46, XX
16	42	23⁺³	Male	Advanced maternal age	46, XY