TY - JOUR

T1 - Multiple mechanisms confining RNA polymerase II ubiquitylation to polymerases undergoing transcriptional arrest

AU - Somesh, Baggavalli P.

AU - Reid, James

AU - Liu, Wei Feng

AU - Søgaard, T. Max M.

AU - Erdjument-Bromage, Hediye

AU - Tempst, Paul

AU - Svejstrup, Jesper Q.

N1 - Funding Information: This work was supported by a grant from Cancer Research UK (to J.Q.S.). We thank Drs. Stefan Jentsch, Daniel Finley, Mike Hampsey, Jack Greenblatt, Roger Kornberg, Jon Huibregtse, and Ron Hay for kind gifts of strains or plasmids. Members of the Svejstrup lab and Drs. Peter Verrijzer and Arnold Kristjuhan are thanked for comments on the manuscript.

PY - 2005/6/17

Y1 - 2005/6/17

N2 - In order to study mechanisms and regulation of RNA polymerase II (RNAPII) ubiquitylation and degradation, highly purified factors were used to reconstitute RNAPII ubiquitylation in vitro. We show that arrested RNAPII elongation complexes are the preferred substrates for ubiquitylation. Accordingly, not only DNA-damage-dependent but also DNA-damage-independent transcriptional arrest results in RNAPII ubiquitylation in vivo. Def1, known to be required for damage-induced degradation of RNAPII, stimulates ubiquitylation of RNAPII only in an elongation complex. Ubiquitylation of RNAPII is dependent on its C-terminal repeat domain (CTD). Moreover, CTD phosphorylation at serine 5, a hallmark of the initiating polymerase, but not at serine 2, a hallmark of the elongating polymerase, completely inhibits ubiquitylation. In agreement with this, ubiquitylated RNAPII is hypophosphorylated at serine 5 in vivo, and mutation of the serine 5 phosphatase SSU72 inhibits RNAPII degradation. These results identify several mechanisms that confine ubiquitylation of RNAPII to the forms of the enzyme that arrest during elongation.

AB - In order to study mechanisms and regulation of RNA polymerase II (RNAPII) ubiquitylation and degradation, highly purified factors were used to reconstitute RNAPII ubiquitylation in vitro. We show that arrested RNAPII elongation complexes are the preferred substrates for ubiquitylation. Accordingly, not only DNA-damage-dependent but also DNA-damage-independent transcriptional arrest results in RNAPII ubiquitylation in vivo. Def1, known to be required for damage-induced degradation of RNAPII, stimulates ubiquitylation of RNAPII only in an elongation complex. Ubiquitylation of RNAPII is dependent on its C-terminal repeat domain (CTD). Moreover, CTD phosphorylation at serine 5, a hallmark of the initiating polymerase, but not at serine 2, a hallmark of the elongating polymerase, completely inhibits ubiquitylation. In agreement with this, ubiquitylated RNAPII is hypophosphorylated at serine 5 in vivo, and mutation of the serine 5 phosphatase SSU72 inhibits RNAPII degradation. These results identify several mechanisms that confine ubiquitylation of RNAPII to the forms of the enzyme that arrest during elongation.

U2 - 10.1016/j.cell.2005.04.010

DO - 10.1016/j.cell.2005.04.010

M3 - Journal article

C2 - 15960978

AN - SCOPUS:20444428382

VL - 121

SP - 913

EP - 923

JO - Cell

JF - Cell

SN - 0092-8674

IS - 6

ER -