Stefin B interacts with histones and cathepsin L in the nucleus.
Stefin B (cystatin B) is an endogenous inhibitor of cysteine proteinases localized in the nucleus and the cytosol. Loss-of-function mutations in the stefin B gene (CSTB) gene were reported in patients with Unverricht-Lundborg disease (EPM1). We have identified an interaction between stefin B and nucleosomes, specifically with histones H2A.Z, H2B, and H3. In synchronized T98G cells, stefin B co-immunoprecipitated with histone H3, predominantly in the G(1) phase of the cell cycle. Stefin B-deficient mouse embryonic fibroblasts entered S phase earlier than wild type mouse embryonic fibroblasts. In contrast, increased expression of stefin B in the nucleus delayed cell cycle progression in T98G cells. The delay in cell cycle progression was associated with the inhibition of cathepsin L in the nucleus, as judged from the decreased cleavage of the CUX1 transcription factor. In vitro, inhibition of cathepsin L by stefin B was potentiated in the presence of histones, whereas histones alone did not affect the cathepsin L activity. Interaction of stefin B with the Met-75 truncated form of cathepsin L in the nucleus was confirmed by fluorescence resonance energy transfer experiments in the living cells. Stefin B could thus play an important role in regulating the proteolytic activity of cathepsin L in the nucleus, protecting substrates such as transcription factors from its proteolytic processing.
Čeru
Slavko
S
Departments of Biochemistry and Molecular and Structural Biology, Ljubljana SI-1000, Slovenia.
Konjar
Špela
Š
Departments of Biochemistry and Molecular and Structural Biology, Ljubljana SI-1000, Slovenia.
Maher
Katarina
K
Departments of Biochemistry and Molecular and Structural Biology, Ljubljana SI-1000, Slovenia.
Repnik
Urška
U
Departments of Biochemistry and Molecular and Structural Biology, Ljubljana SI-1000, Slovenia.
Križaj
Igor
I
Molecular and Biomedical Sciences, Jožef Stefan Institute, Ljubljana SI-1000, Slovenia.
Benčina
Mojca
M
Department of Biotechnology, National Institute of Chemistry, Ljubljana SI-1000, Slovenia.
Renko
Miha
M
Departments of Biochemistry and Molecular and Structural Biology, Ljubljana SI-1000, Slovenia.
Nepveu
Alain
A
Molecular Oncology Group, McGill University, Montreal, Quebec H3A 1A1, Canada.
Žerovnik
Eva
E
Departments of Biochemistry and Molecular and Structural Biology, Ljubljana SI-1000, Slovenia.
Turk
Boris
B
Departments of Biochemistry and Molecular and Structural Biology, Ljubljana SI-1000, Slovenia.
Kopitar-Jerala
Nataša
N
Departments of Biochemistry and Molecular and Structural Biology, Ljubljana SI-1000, Slovenia. Electronic address: natasa.kopitar@ijs.si.
D000818:Animals; D056668:Cathepsin L / Q000378:metabolism*; D002453:Cell Cycle; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism*; D055313:Cystatin B / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D005786:Gene Expression Regulation; D006657:Histones / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D051379:Mice; D008954:Models, Biological
D006657:Histones; D055313:Cystatin B; D056668:Cathepsin L