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JMJD6-STAT3^Y705ph axis promotes autophagy in osteosarcoma cancer cells by regulating ATG

Kun Wang^1,2, Baoyin Du¹, Bin Xu¹

¹Department of Emergency and Trauma Surgery, Jining No.1 People's Hospital, Jining, China.

Biofactors (Oxford, England)

|January 21, 2020

Abstract

Background:

Osteosarcoma cancer is a malignant tumor with poor outcome. Activation of STAT3 is closely related with tumor development. We intended to study the effects of JMJD6 on phosphorylation of STAT3 at Y705 site.

Methods:

Osteosarcoma cancer cell lines (Saos-2, MG-63, and HOS) and clinical specimens were obtained. Interference RNA or JMJD6 mimic was transfected into the cells to silence or mimic JMJD6. Immunoprecipitation assay and glutathione S-transferase (GST) pull down was implemented to investigate whether JMJD6 is associated with STAT3. STAT3-null HOS cells were simultaneously transfected with the plasmids bearing STAT3^WT or STAT3^Y705F and EGFP-tagged LC3 plasmids. Recombinant full-length JMJD6 protein was subjected to in vitro kinase activity assay for testing its ability to phosphorylate STAT3. The severity of autophagy was indicated by the number of autophagosomes, expression of EGFP-LC3, ratio of LC3-II to LC3-I, degradation percentage of long-lived proteins and expression of autophagy associated gene (ATG).

Results:

JMJD6 modulated the phosphorylation of STAT3 at Y705 site in osteosarcoma cells. Results from immunoprecipitation and GST pull down assays showed that JMJD6 associated with STAT3 in osteosarcoma cells. JMJD6 silence impeded the formation of autophagosomes, inhibited the accumulation of EGFP-LC3, decreased the ratio of LC3-II to LC3-I, blocked the degradation of long-lived proteins, and repressed the expression of ATG. JMJD6-induced autophagy was impaired by STAT3^Y705F which was not phosphorylated by JMJD6. The JMJD6-STAT3^Y705ph axis was implicated in the transcriptional regulation of ATG.

Conclusion:

JMJD6 was included in regulating the phosphorylation of STAT3^Y705 and promoted autophagy of osteosarcoma cells through its kinase activity.

Insights

JMJD6 regulates STAT3 phosphorylation at Y705, promoting osteosarcoma cell autophagy. This kinase activity is crucial for tumor development and may offer therapeutic targets.

Area of Science:

Oncology
Molecular Biology
Cell Biology

Background:

Osteosarcoma is a challenging cancer with poor prognosis.
Signal transducer and activator of transcription 3 (STAT3) activation is linked to tumor progression.
The role of Jumonji domain-containing protein 6 (JMJD6) in osteosarcoma tumorigenesis requires further investigation.

Purpose of the Study:

To investigate the effect of JMJD6 on STAT3 phosphorylation at the Y705 site in osteosarcoma.
To elucidate the mechanism by which JMJD6 influences osteosarcoma cell autophagy.
To determine the functional relationship between JMJD6, STAT3, and autophagy in osteosarcoma.

Main Methods:

Utilized osteosarcoma cell lines and clinical specimens.
Employed RNA interference and mimic transfection to modulate JMJD6 expression.
Performed immunoprecipitation, GST pull-down, and in vitro kinase assays to assess protein interactions and activity.
Analyzed autophagy markers including autophagosome formation, LC3 conversion, and protein degradation.

Main Results:

JMJD6 directly modulates STAT3 phosphorylation at the Y705 site in osteosarcoma cells.
JMJD6 physically associates with STAT3.
JMJD6 promotes osteosarcoma cell autophagy, evidenced by increased autophagosome formation and ATG gene expression.
Impaired STAT3 phosphorylation at Y705 (STAT3^Y705F) abrogated JMJD6-induced autophagy.
The JMJD6-STAT3^Y705ph pathway regulates the transcription of autophagy-associated genes (ATG).

Conclusions:

JMJD6 acts as a kinase, phosphorylating STAT3 at Y705.
JMJD6 promotes osteosarcoma cell autophagy through its kinase activity on STAT3.
The JMJD6-STAT3 axis is a key regulator of autophagy in osteosarcoma, suggesting potential therapeutic implications.

Read full analysis

Keywords:

ATG JMJD6 STAT3Y705ph autophagy osteosarcoma

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