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CDK1‑induced regulation of p53 phosphorylation at Ser315 mediates cell cycle arrest and apoptosis of macrophages infected with clinical isolates of Mycobacterium tuberculosis

  • Authors:
    • Banghao Sun
    • Zhenyu Zhao
    • Qingyu Meng
    • Luya Pu
    • Xingyu Jiang
    • Shuai Li
    • Fadi Cao
    • Fan Li
  • View Affiliations / Copyright

    Affiliations: Department of Immunology, School of Basic Medical Sciences, Xinjiang Medical University, Urumqi, Xinjiang 830000, P.R. China, School and Hospital of Stomatology, Jilin University, Changchun, Jilin 130021, P.R. China, Department of Pathogen Biology, The Key Laboratory of Zoonosis, Chinese Ministry of Education, College of Basic Medicine, Jilin University, Changchun, Jilin 130021, P.R. China
    Copyright: © Sun et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 44
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    Published online on: November 20, 2025
       https://doi.org/10.3892/mmr.2025.13754
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Abstract

Tuberculosis (TB) is an infectious disease caused by infection with Mycobacterium tuberculosis (MTB). The morbidity of TB in the Xinjiang region of China is higher than that in other provinces. Macrophage apoptosis after infection with MTB is considered to serve a key role in killing the bacteria. However, the biological process of apoptosis and the underlying molecular mechanisms triggered by the infection of macrophages with clinical isolates of MTB from Xinjiang (XJMTB) are not clear. The present study aimed to investigate the unique characteristics of XJMTB. Briefly, western blotting and flow cytometry were employed in the present study, and it was demonstrated that macrophages infected with MTB H37Rv or XJMTB underwent G2/M cell cycle arrest and apoptosis. The transcriptome sequencing analysis showed that cyclin‑dependent kinase 1 (CDK1) was a key regulatory gene in regulating the G2/M cell cycle arrest and apoptosis in MTB‑infected macrophages, and the p53 gene was most likely involved in the regulation of this. Moreover, the phosphorylation of p53 (Ser315) was elevated with the upregulation of CDK1 activation, leading to a higher proportion of MTB‑infected macrophages exhibiting G2/M cell cycle block and apoptosis. The current study also revealed that enhanced activation of CDK1 reversed the attenuation of the G2/M cell cycle block and the reduction in the percentage of apoptosis caused by inhibition of p53 (Ser315) phosphorylation. Furthermore, the co‑immunoprecipitation experiment demonstrated an interaction between CDK1 and p53. The present study indicated that, in an in vitro model of macrophage infection with XJMTB, enhanced activation of CDK1 may regulate the phosphorylation of p53 (Ser315), promote the secretion of TNF‑α, IL‑6, IL‑10, IL‑1β and IL‑12, promote G2/M cell cycle arrest and apoptosis of macrophages, and enhance the survival of XJMTB in macrophages. These results provide CDK1 and phosphorylated‑p53 as two new potential therapeutic targets for TB in Xinjiang, and lay a foundation for the development of novel TB treatment strategies.
View Figures

Figure 1

Isolation and acid-fast staining of
Mycobacterium tuberculosis. (A) Results of isolation in
modified acidic Roche's medium. (B) Acid-fast staining of an
isolated Mycobacterium tuberculosis from Xinjang strain.

Figure 2

Secretion levels of cytokines at
different time points after MTB infection of macrophages and the
secretion level of NO 24 h after infection. Levels of (A) TNF-α,
(B) IL-1β, (C) IL-12 (p70), (D) IL-10 and (E) IL-6 secretion at
different time points after MTB infection in macrophages. (F)
Levels of NO secretion in macrophages infected with MTB for 24 h.
Data are presented as the mean ± SEM. *P<0.05, **P<0.01,
***P<0.001. C group, blank control group not infected with any
bacteria; L group, MTB from Xinjang clinical isolate-infected
group; MTB, Mycobacterium tuberculosis; NO, nitric oxide;
ns, not significant; V group, H37Rv standard strain-infected
group.

Figure 3

CDK1 expression is upregulated in
response to infection with MTB. Heatmap of DEGs in (A) L and (B) V
groups vs. C group. Red, high expression; blue, low expression. (C)
Principal component analysis of group L, group V and group C.
Volcano plots of DEGs in the (D) L and (E) V groups (|log2 fold
change|≥1; P<0.05). Green, downregulated expression; gray, no
differential expression; red, upregulated expression. (F) Venn
diagram of L vs. C DEGs and V vs. C DEGs. GO functional enrichment
analysis of 1,527 mRNAs: (G) Cellular components, (H) biological
processes and (I) molecular functions. (J) Kyoto Encyclopedia of
Genes and Genomes pathway enrichment analysis of 1,527 mRNAs. (K)
Protein-protein interaction network top-level module containing 35
genes modularized by the plug-in MCODE. GO functional enrichment
analysis of 35 key mRNAs: (L) Biological processes, (M) cellular
components and (N) molecular functions. (O) Top 30 genes of the
1,527 mRNAs contained in the module that interacted most closely
with other genes in the module. Expression of (P) CDK1 and (Q)
CCNA2 in each group was verified through reverse
transcription-quantitative polymerase chain reaction. Data are
presented as the mean ± SEM. ***P<0.001. C group, blank control
group not infected with any bacteria; CCNA2, cyclin A 2; CDK1,
cyclin-dependent kinase; DEG, differentially expressed gene; GO,
Gene Ontology; L group, MTB from Xinjang clinical isolate-infected
group; MTB, Mycobacterium tuberculosis; V group, H37Rv
standard strain-infected group.

Figure 4

Infection with XJMTB causes stronger
G2/M cell cycle arrest and apoptosis than infection with
H37Rv. Cell cycle profile of macrophages in the (A) C group, (B) V
group, (C) L3 group, (D) L4 group and (E) L5 group. (F)
G2/M cell cycle ratio in each group. (G) Protein
expression levels of BCL2, BAX, BCL-2, caspase 3 and
cleaved-caspase 3 were determined by western blotting. (H)
Statistical analysis of apoptosis-related proteins detected by
western blotting. Apoptosis of macrophages in the (I) C group, (J)
V group, (K) L3 group, (L) L4 group and (M) L5 group. (N) Apoptosis
rate in each group. Data are presented as the mean ± SEM.
**P<0.01, ***P<0.001. A.U., arbitrary units; C group, blank
control group not infected with any bacteria; L group, XJMTB
clinical isolate-infected group; MTB, Mycobacterium
tuberculosis; V group, H37Rv standard strain-infected group;
XJMTB, MTB from Xinjang.

Figure 5

Inhibition of CDK1 alleviates
macrophage G2/M cycle arrest and apoptosis caused by MTB
infection. (A) Protein expression levels of CDK1 were determined by
western blotting. (B) Statistical analysis of CDK1 protein
expression detected by western blotting. Cell cycle profiles of
macrophages in the (C) V group, (D) L3 group, (E) L4 group and (F)
L5 group when Ro3306 was not added. Cell cycle profiles of
macrophages in the (G) V group, (H) L3 group, (I) L4 group and (J)
L5 group after the addition of Ro3306. Apoptosis of macrophages in
the (K) V group, (L) L3 group, (M) L4 group and (N) L5 group when
Ro33006 was not added. Apoptosis of macrophages in the (O) V group,
(P) L3 group, (Q) L4 group and (R) L5 group after the addition of
Ro3306. (S) G2/M cell cycle ratio and (T) apoptosis rate
in each group before and after the addition of Ro3306. (U) Protein
expression levels of BCL2, BAX, Bcl-2, caspase 3 and
cleaved-caspase 3 were determined by western blotting before and
after the addition of Ro3306. (V) Statistical analysis of
apoptosis-related protein expression detected by western blotting
before and after the addition of Ro3306. Data are presented as the
mean ± SEM. *P<0.05, **P<0.01, ***P<0.001. A.U., arbitrary
units; CDK1, cyclin-dependent kinase; L group, MTB from Xinjang
clinical isolate-infected group; MTB, Mycobacterium
tuberculosis; V group, H37Rv standard strain-infected
group.

Figure 6

Inhibition of p53 (Ser315)
phosphorylation attenuates macrophage G2/M cycle block
and apoptosis caused by MTB infection. (A) Protein expression
levels of total p53 and p-p53 (Ser315) after MTB infection were
determined by western blotting. (B) Statistical analysis of total
p53 and p-p53 (Ser315) expression detected by western blotting
after MTB infection. (C) Protein expression levels of total p53 and
p-p53 (Ser315) were determined by western blotting before and after
the addition of Pft-α. (D) Statistical analysis of total p53 and
p-p53 (Ser315) expression determined by western blotting before and
after the addition of Pft-α. Cell cycle profiles of macrophages in
the (E) V group, (F) L3 group, (G) L4 group and (H) L5 group when
Pift-α was not added. Cell cycle profiles of macrophages in the (I)
V group, (J) L3 group, (K) L4 group and (L) L5 group after the
addition of Pft-α. Apoptosis of macrophages in the (M) V group, (N)
L3 group, (O) L4 group and (P) L5 group when Pft-α was not added.
Apoptosis of macrophages in the (Q), V group, (R) L3 group, (S) L4
group and (T) L5 group after the addition of Pft-α. (U)
G2/M cell cycle ratio and (V) apoptosis rate of each
group before and after the addition of Pft-α. (W) Protein
expression levels of BCL2, BAX, caspase 3 and cleaved-caspase 3
were determined by western blotting before and after the addition
of Pft-α. (X) Statistical analysis of apoptosis-related protein
expression detected by western blotting before and after the
addition of Pft-α. Data are presented as the mean ± SEM.
*P<0.05, **P<0.01, ***P<0.001. A.U., arbitrary units; L
group, MTB from Xinjang clinical isolate-infected group; MTB,
Mycobacterium tuberculosis; n.s., not significant; p-,
phosphorylated; Pft-α, pifithrin-α; V group, H37Rv standard
strain-infected group.

Figure 7

Secretion of cytokines and NO before
and after inhibition of CDK1 expression or inhibition of p53
(Ser315) phosphorylation. Secretion of (A) IL-6, (B) TNF-α, (C)
IL-1β, (D) IL-10, (E) IL-12 and (F) NO before and after inhibition
of CDK1 expression or inhibition of p53 (Ser315) phosphorylation.
Data are presented as the mean ± SEM. ***P<0.001. MTB,
Mycobacterium tuberculosis; NO, nitric oxide; ns, not
significant; Pft-α, pifithrin-α; XJMTB, MTB from Xinjang.

Figure 8

CDK1 mediates G2/M cell
cycle arrest and apoptosis caused by infection with XJMTB through
regulation of p53 (Ser315) phosphorylation. (A) Phosphorylation of
p53 (Ser315) after inhibition of CDK1. (B) Phosphorylation of CDK1
(Thr161) and of p53 (Ser315) following treatment with TC11. (C)
Expression of CDK1 after inhibition of p53 (Ser315)
phosphorylation. (D) Statistical analysis of total p53 and p-p53
(Ser315) expression determined by western blotting before and after
the addition of Ro3306. (E) Statistical analysis of total p53,
p-p53 (Ser315), total CDK1 and p-CDK1 (Thr161) expression
determined by western blotting before and after the addition of
TC11. (F) Statistical analysis of total p53, p-p53 (Ser315) and
total CDK1 expressiondetermined by western blotting before and
after the addition of Pft-α. Cell cycle progression of
XJMTB-infected macrophages in the (G) TC11, (H) Pft-α and (I) TC11
+ Pft-α groups. (J) G2/M cell cycle ratio in the various
groups. Apoptosis of XJMTB-infected macrophages in the (K) TC11,
(L) Pft-α and (M) TC11 + Pft-α groups. (N) Apoptosis rate in the
various groups. (O and P) Protein expression levels of BCL2, BAX,
caspase 3 and cleaved-caspase 3 were determined by western blotting
in the TC11, Pft-α and TC11 + Pft-α groups. (Q) CFU analysis was
performed to determine the survival rate of XJMTB in macrophages in
the TC11, Pft-α and TC11 + Pft-α groups. (R) Co-IP results of CDK1
and p53. Data are presented as the mean ± SEM. *P<0.05,
**P<0.01, ***P<0.001. A.U., arbitrary units; CDK1,
cyclin-dependent kinase; CFU, colony-forming unit; IB,
immunoblotting; IgG, immunoglobulin G; IP, immunoprecipitation; L
group, XJMTB clinical isolate-infected group; MTB, Mycobacterium
tuberculosis; n.s., not significant; p-, phosphorylated; Pft-α,
pifithrin-α; XJMTB, MTB from Xinjang.
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Copy and paste a formatted citation
Spandidos Publications style
Sun B, Zhao Z, Meng Q, Pu L, Jiang X, Li S, Cao F and Li F: CDK1‑induced regulation of p53 phosphorylation at Ser315 mediates cell cycle arrest and apoptosis of macrophages infected with clinical isolates of <em>Mycobacterium tuberculosis</em>. Mol Med Rep 33: 44, 2026.
APA
Sun, B., Zhao, Z., Meng, Q., Pu, L., Jiang, X., Li, S. ... Li, F. (2026). CDK1‑induced regulation of p53 phosphorylation at Ser315 mediates cell cycle arrest and apoptosis of macrophages infected with clinical isolates of <em>Mycobacterium tuberculosis</em>. Molecular Medicine Reports, 33, 44. https://doi.org/10.3892/mmr.2025.13754
MLA
Sun, B., Zhao, Z., Meng, Q., Pu, L., Jiang, X., Li, S., Cao, F., Li, F."CDK1‑induced regulation of p53 phosphorylation at Ser315 mediates cell cycle arrest and apoptosis of macrophages infected with clinical isolates of <em>Mycobacterium tuberculosis</em>". Molecular Medicine Reports 33.1 (2026): 44.
Chicago
Sun, B., Zhao, Z., Meng, Q., Pu, L., Jiang, X., Li, S., Cao, F., Li, F."CDK1‑induced regulation of p53 phosphorylation at Ser315 mediates cell cycle arrest and apoptosis of macrophages infected with clinical isolates of <em>Mycobacterium tuberculosis</em>". Molecular Medicine Reports 33, no. 1 (2026): 44. https://doi.org/10.3892/mmr.2025.13754
Copy and paste a formatted citation
x
Spandidos Publications style
Sun B, Zhao Z, Meng Q, Pu L, Jiang X, Li S, Cao F and Li F: CDK1‑induced regulation of p53 phosphorylation at Ser315 mediates cell cycle arrest and apoptosis of macrophages infected with clinical isolates of <em>Mycobacterium tuberculosis</em>. Mol Med Rep 33: 44, 2026.
APA
Sun, B., Zhao, Z., Meng, Q., Pu, L., Jiang, X., Li, S. ... Li, F. (2026). CDK1‑induced regulation of p53 phosphorylation at Ser315 mediates cell cycle arrest and apoptosis of macrophages infected with clinical isolates of <em>Mycobacterium tuberculosis</em>. Molecular Medicine Reports, 33, 44. https://doi.org/10.3892/mmr.2025.13754
MLA
Sun, B., Zhao, Z., Meng, Q., Pu, L., Jiang, X., Li, S., Cao, F., Li, F."CDK1‑induced regulation of p53 phosphorylation at Ser315 mediates cell cycle arrest and apoptosis of macrophages infected with clinical isolates of <em>Mycobacterium tuberculosis</em>". Molecular Medicine Reports 33.1 (2026): 44.
Chicago
Sun, B., Zhao, Z., Meng, Q., Pu, L., Jiang, X., Li, S., Cao, F., Li, F."CDK1‑induced regulation of p53 phosphorylation at Ser315 mediates cell cycle arrest and apoptosis of macrophages infected with clinical isolates of <em>Mycobacterium tuberculosis</em>". Molecular Medicine Reports 33, no. 1 (2026): 44. https://doi.org/10.3892/mmr.2025.13754
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