Page 15 - 南京医科大学自然版

第46卷第10期 王新新，黄伯贤，陆嘉逢. PUS7通过mRNA假尿嘧啶化调控胚胎发育过程中神经
2026年10月 谱系分化平衡［J］. 南京医科大学学报（自然科学版），2026，46（10）：1437-1449 ·1445 ·

［9］
或表达维持 ，这类同步变化的基因更可能反映 联。因此，本研究重点关注Ψ修饰下降且mRNA表达
Pus7缺失后Ψ修饰降低与转录本表达受损之间的关 下调的交集基因，以筛选可能受PUS7介导mRNA Ψ
A d 0 d 2 d 3 d 5 B WT Pus7⁃KO

N2B27 N2B27+RA
d 1
Neuronal differentiation
Stage 1 Stage 2
C Pluripotency Neuroectoderm⁃related genes
d 3
d 0
log2 FC
（KO/WT）
d 3 2
0
Klf4 Sox2 Pou5f1 Nanog Ncam1 Sox9 Map2k6 Map2 En2 Nptx2 Hes5 Aldh1l1 Otx1 Serf1 Nr2f1 Meis1 Sox3 Edn3 Irx2 Mkx6⁃1 Meis2 Gli3 Prdm1 Meis3 Pax2 Phox2b Nsg2 Zic5 En1 Pax6 Six3 Sox1 Pax3 Olig2 -2 d 5

D WT WT WT WT WT
Pus7⁃KO Pus7⁃KO Pus7⁃KO Pus7⁃KO Pus7⁃KO
40 25 15 8 *** 1.5
***
mRNA level 30 Sox1 mRNA level 15 mRNA level 10 ** *** mRNA level 6 4 Aldh1l1 mRNA level 1.0
20
***
20
Pax6 10 10 5 ** Map2 5 Ncam1 2 * 0.5

0 0 0 0 0
d0 d3 d5 d0 d3 d5 d0 d3 d5 d0 d3 d5 d0 d3 d5
E F G
WT Pus7⁃KO Pus7⁃KO WT 3.0×10 3
WT
d0 d3 d5 d0 d3 d5 Merge SOX1 Merge SOX1
Pus7⁃KO
PAX6 50 kDa 2.0×10 3
Count
GAPDH 36 kDa 0
d 3
1.0×10
PAX6 * WT 0 3 3 4 5
protein level 40 Pus7⁃KO 3 d H -10 0 10 10 10
**
SOX1⁃FITC
30
20
***
PAX6 10 （of DAPI + ） 50
40
0
d0 d3 d5 5 cells 30
20
d
% FITC + 10 0
WT Pus7⁃KO
A：Schematic illustration of the experimental workflow for directed neural ectoderm differentiation of mESC. B：Representative bright⁃field images of
WT and Pus7⁃KO cells at the indicated time points during directed neural ectoderm differentiation（Scale bar=100 μm）. C：Heatmap showing the log 2 FC in
the expression of pluripotency⁃ and neuroectoderm⁃associated marker genes in Pus7⁃KO relative to WT at day 0 and day 3 of directed neural ectoderm
differentiation（n=3）. D：qRT⁃PCR analysis of the mRNA expression of neural progenitor cellmarker genes（Sox1 and Pax6），mature neuronal marker genes
（Ncam1 and Map2）and glial marker gene（Aldh1l1）in WT and Pus7⁃KO cells during directed neural ectoderm differentiation（n=3）. E：Western blot anal⁃
ysis of PAX6 protein expression in WT and Pus7⁃KO cells during directed neural ectoderm differentiation（n=3）. Quantification of PAX6 protein levels is
shown on the right. F：Immunofluorescence staining of SOX1 in WT and Pus7⁃KO cells during directed neural ectoderm differentiation；DAPI was used for
nuclear staining（blue），and SOX1 is shown in green. Scale bar=50 μm（n=3）. G：Distribution of SOX1⁃FITC fluorescence signals in WT and Pus7⁃KO cells
***
**
*
at d3（n=3）. H：Quantification of the proportion of SOX1⁃positive cells in WT and Pus7⁃KO groups. P < 0.05，P < 0.01，and P < 0.001.
图4 Pus7敲除促进早期神经祖细胞程序增强并伴随后续成熟相关程序受阻
Figure 4 Pus7 knockout enhanced early neural progenitor cell programs while impairing subsequent maturation⁃related
programs