Related Experiment Videos
Identification of a nuclear Stat1 protein tyrosine phosphatase
Johanna ten Hoeve1, Maria de Jesus Ibarra-Sanchez, Yubin Fu
1Department of Medicine, University of California-Los Angeles, Los Angeles, California 90095, USA.
Abstract:
Upon interferon (IFN) stimulation, Stat1 becomes tyrosine phosphorylated and translocates into the nucleus, where it binds to DNA to activate transcription. The activity of Stat1 is dependent on tyrosine phosphorylation, and its inactivation in the nucleus is accomplished by a previously unknown protein tyrosine phosphatase (PTP). We have now purified a Stat1 PTP activity from HeLa cell nuclear extract and identified it as TC45, the nuclear isoform of the T-cell PTP (TC-PTP). TC45 can dephosphorylate Stat1 both in vitro and in vivo. Nuclear extracts lacking TC45 fail to dephosphorylate Stat1. Furthermore, the dephosphorylation of IFN-induced tyrosine-phosphorylated Stat1 is defective in TC-PTP-null mouse embryonic fibroblasts (MEFs) and primary thymocytes. Reconstitution of TC-PTP-null MEFs with TC45, but not the endoplasmic reticulum (ER)-associated isoform TC48, rescues the defect in Stat1 dephosphorylation. The dephosphorylation of Stat3, but not Stat5 or Stat6, is also affected in TC-PTP-null cells. Our results identify TC45 as a PTP responsible for the dephosphorylation of Stat1 in the nucleus.
Insights
A novel protein tyrosine phosphatase (PTP), TC45, has been identified as the key enzyme responsible for dephosphorylating Signal transducer and activator of transcription 1 (Stat1) in the cell nucleus, regulating its activity after interferon stimulation.
Area of Science:
- Molecular Biology
- Cell Biology
- Immunology
Background:
- Signal transducer and activator of transcription 1 (Stat1) is crucial for interferon-mediated gene transcription.
- Stat1 activity is regulated by tyrosine phosphorylation and subsequent dephosphorylation.
- The nuclear protein tyrosine phosphatase (PTP) responsible for Stat1 inactivation was previously unknown.
Purpose of the Study:
- To identify the nuclear PTP responsible for Stat1 dephosphorylation.
- To characterize the role of this PTP in Stat1 regulation.
- To investigate the isoform specificity of PTP activity on Stat1.
Main Methods:
- Purification of Stat1 PTP activity from HeLa cell nuclear extracts.
- In vitro and in vivo dephosphorylation assays using Stat1.
- Analysis of Stat1 dephosphorylation in TC-PTP-null mouse embryonic fibroblasts (MEFs) and thymocytes.
- Reconstitution experiments with TC45 and TC48 isoforms.
Main Results:
- A Stat1 PTP activity was purified and identified as TC45, the nuclear isoform of T-cell PTP (TC-PTP).
- TC45 dephosphorylates Stat1 both in vitro and in vivo.
- Stat1 dephosphorylation is impaired in TC-PTP-null cells and can be rescued by TC45, but not TC48.
- TC45 also affects Stat3 dephosphorylation, but not Stat5 or Stat6.
Conclusions:
- TC45 is the primary nuclear PTP responsible for dephosphorylating Stat1.
- This finding elucidates a key mechanism for regulating Stat1 activity in the nucleus.
- Isoform specificity of TC-PTP is critical for its function in Stat1 dephosphorylation.