Bethyl Laboratories, Inc.
Antibodies to 
Integrator Complex Proteins
March 9, 2010
Overview

The integrator complex was identified in a study of DSS1 (deleted in split hand/split foot 1), a factor that associates with BRCA2 and the 19S proteasome (1). The core integrator complex is a 12 subunit complex (INTS1-INTS12) that associates with the C-terminal domain of the largest subunit of the RNA polymerase II (RNAPII) holoenzyme.  In association with RNAPII, the integrator complex mediates the 3’ end processing of small nuclear RNAs U1 and U2.  Recently, additional components of the integrator core complex have been identified (2).  New members of the extended integrator complex include a phosphatase module (PPP1CB, PPP2CA/B, and PPP2R1A/B), four uncharacterized proteins (C12orf11, C15orf44, C7orf26, and C9orf80), two OB-fold proteins (OBFC2A and OBFC2B), and a set of zinc-finger proteins (ZMYND8, ZNF687, and ZNF589). The functions of some of the integrator subunits have been characterized and linked to a variety of cellular processes and signaling pathways.  INTS11 is the catalytic subunit of the integrator core complex and is directly involved in the processing of U1 and U2 snRNA genes (1). INTS6/DICE1 has been implicated to play a role in Wnt signaling (3), and INTS3 has been shown to associate in a complex with human single-stranded binding protein (hSSB1) and possibly play a role in the DNA damage response independent of the integrator core complex (4).

References
1. D. Baillat et al., "Integrator, a Multiprotein Mediator of Small Nuclear RNA Processing, Associates With the C-Terminal Repeat of RNA Polymerase II," Cell. 123, no. 2 (2005): 265-276.
2. A. Malovannaya et al., "Streamlined Analysis Schema for High-Throughput Identification of Endogenous Protein Complexes," Proc.Natl.Acad.Sci U.S.A. 107, no. 6 (2010): 2431-2436.
3. S. Filleur et al., "INTS6/DICE1 Inhibits Growth of Human Androgen-Independent Prostate Cancer Cells by Altering the Cell Cycle Profile and Wnt Signaling," Cancer Cell Int. 9 (2009): 28.
4. J. R. Skaar et al., "INTS3 Controls the HSSB1-Mediated DNA Damage Response," J Cell Biol. 187, no. 1 (2009): 25-32.

Bethyl Laboratories' Portfolio
INT1
INT3 (FLJ21919)
INT4
INT5
INT6 (DDX26/DICE1)
INT7 (DKFZP434B168)
INT8 (FLJ20530/KAONASHI1)
INTS9 (RC74)
PP1CB
PPP2R1A
OBFC2A
SSB1
RNA Polymeriase II



Detection of Human INT6 by WB and IP.

Samples: Whole cell lysate  (5, 15 and 50 mcg for WB; 1 mg for IP, 20% of IP loaded) from HeLa cells.  Antibodies: Affinity purified rabbit anti-INT6 antibody A301-658A used for WB at 0.04 mcg/ml (A) and 1 mcg/ml (B) and used for IP at 3 mcg/mg lysate. INT6 was also immunoprecipitated by rabbit anti-INT6 antibody BL6264 which recognizes an upstream epitope.  For blotting immunoprecipitated INT6, ReliaBLOT Reagents and Procedures (Cat. No. WB120) were used.  Detection: Chemiluminescence with exposure times of 10 seconds (A) and 1 second (B).

Detection of Phospho RNA Polymerase II (S5) in Whole Cell Lysates and IP.

Samples: Whole cell lysate (50 mcg for A and Input for B; 1 mg for IP, 20% of IP loaded) from HeLa cells.  In B, immunoprecipitates were either mock treated or treated with lambda phosphatase.  Antibodies: Affinity purified Phospho RNA Polymerase II (S5) antibody BL2896 (Cat. No. A300-655A) used for WB at 0.02 mcg/ml (A) and 0.1 mcg/ml (B). In A, BL2896 was pre-incubated without peptide (-), with non-phosphorylated peptide (NP), with phosphorylated S2 peptide, or with phosphorylated S5 peptide.  In B, RNA Polymerase II was immunoprecipitated using RNA Polymerase II antibody BL2893 (Cat. No. A300-653A) at 3 mcg/mg lysate.  Immunoprecipitates were either mock treated or treated with lambda phosphatase. For blotting immunoprecipitated RNA Polymerase II, ReliaBLOT Reagents and Procedures (Cat. No. WB120) were used.  Detection: Chemiluminescence with exposure times of 10 seconds.
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Bethyl Laboratories' portfolio of polyclonal antibodies commercially available includes over 3500 polyclonal antibodies to over 1300 proteins. We test our antibodies in a wide range of applications including ELISA, western blot, immunoprecipitation, immunohistochemistry, and immunocytochemistry. We have recently validated many of our antibodies for flow cytometry and ChIP. Our antibody range also includes secondary antibodies, loading controls and antibodies to epitope tags.
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