Bruce Knutson, PhD

RESEARCH ABSTRACT

Fig.1. The nucleolus

A century old hallmark of cancer is an enlarged nucleolus (Fig.1), a unique nuclear sub compartment where RNA polymerase I (Pol I) transcription and ribosome biogenesis take place. Pol I transcription is unregulated in cancer cells and drives cell proliferation, making it an attractive anti-cancer therapeutic target. The major focus of our research is to elucidate the molecular mechanism of Pol I transcription and how its dysregulation leads to cancer and disease. Our research uses an innovative cross-organismal and interdisciplinary approach that integrates bioinformatics, biochemistry, computational biology, genetics, proteomics and structural biology in yeast and human model system.

Molecular architecture of the Pol I preinitiation complex (PIC)

Fig. 2. Pol I PIC Model

Pol I transcription begins with the formation of the PIC (Fig.2), a macromolecular assemblage of more than 20 different proteins that function coordinately to accurately position Pol I at the promoter and to help initiate transcription. We are interested in the key structural facets of Pol I PIC formation and how it's altered in cancer and diseased cells. Our lab uses an integrated combination of sophisticated protein-protein interaction mapping technologies such as combined chemical crosslinking/mass spectrometry to determine the spatial orientation of Pol I PIC components and how they change during the transcription cycle and in diseased states.

Pol I and craniofacial dysmorphology. Mutations in Pol I cause an autosomal dominant craniofacial abnormality called Treacher Collins Syndrome (TCS). TCS is characteried by an underdeveloped lower jaw and cheekbones that is treated by an extensive multi-stage surgical reconstruction from childhood to early adulthood. We are interested in how these Pol I mutations cause TCS, how they affect Pol I activity, and how they can be suppressed to prevent the disease. Currently, there are no known cures for TCS and other related craniofacial dysmorphologies.

Fig.3. Pol I Regulation

Pol I dysregulation in cancer. The upregulation of Pol I transcription in cancer cells coincides with activating mutations in many oncogenes and loss of function mutations in tumor suppressors that are believed to directly regulate Pol I activity (Fig.3). However, their bona fide Pol I targets and sites of interaction remain unclear. To understand how these cancer proteins target the Pol I complex, we use a combination of protein crosslinking technologies coupled of molecular genetics and biochemistry to identify and characterize the direct and functionally relevant in vivo Pol I targets. These studies will illuminate new strategies to control aberrant Pol I activity.

Graduate research in the Knutson Lab. Interested students should directly contact Bruce Knutson to discuss available research opportunities.

Knutson BA, McNamar R, Rothblum LI. Dynamics of the RNA polymerase I TFIIF/TFIIE-like subcomplex: a mini-review. Biochem Soc Trans. 2020 Oct 30;48(5):1917-1927. doi: 10.1042/BST20190848.

Knutson BA, Smith ML, Belkevich AE, Fakhouri AM. Molecular Topology of RNA Polymerase I Upstream Activation Factor. Mol Cell Biol. 2020 Jun 15;40(13):e00056-20. doi: 10.1128/MCB.00056-20. Print 2020 Jun 15.

McNamar R, Abu-Adas Z, Rothblum K, Knutson BA, Rothblum LI. Conditional depletion of the RNA polymerase I subunit PAF53 reveals that it is essential for mitosis and enables identification of functional domains. J Biol Chem. 2019 Dec 27;294(52):19907-19922. doi: 10.1074/jbc.RA119.009902. Epub 2019 Nov

Jackobel AJ, Zeberl BJ, Glover DM, Fakhouri AM, Knutson BA. DNA binding preferences of S. cerevisiae RNA polymerase I Core Factor reveal a preference for the GC-minor groove and a conserved binding mechanism. Biochim Biophys Acta Gene Regul Mech. 2019 Sep;1862(9):194408. doi: 10.1016/j.bbagrm.2019.194408. Epub 2019 Aug 2.

Smith ML, Cui W, Jackobel AJ, Walker-Kopp N, Knutson. Reconstitution of RNA Polymerase I Upstream Activating Factor and the Roles of Histones H3 and H4 in Complex Assembly. J Mol Biol. 2018 Epub

Jackobel AJ, Han Y, He Y, Knutson BA. Breaking the mold: structures of the RNA polymerase I transcription complex reveal a new path for initiation. Transcription. 2018 Jan 15:1-7

Walker-Kopp N, Jackobel AJ, Pannafino GN, Morocho PA, Xu X, Knutson BA. Treacher Collins syndrome mutations in Saccharomyces cerevisiae destabilize RNA polymerase I and III complex integrity. Hum Mol Genet. 2017 Nov 1;26(21):4290-4300.

Han Y, Yan C, Nguyen THD, Jackobel AJ, Ivanov I, Knutson BA, He Y. Structural mechanism of ATP-independent transcription initiation by RNA polymerase I. Elife. 2017 Jun 17;6.

Knutson BA, Smith ML, Walker-Kopp N, Xu X. Super elongation complex contains a TFIIF-related subcomplex. Transcription. 2016. 7(4):133-40

Knutson BA, Lui J, Ranish J. Hahn S. Architecture of the S.cerevisiae RNA polymerase I Core Factor complex. Nature Structural and Molecular Biology. 2014. 21(9): 810-816

Knutson BA, Hahn S. TFIIB-related factors in RNA polymerase I transcription. Biochem Biophys Acta. 2013. 1829(3-4): 265-273

Knutson BA. Emergence and expansion of TFIIB-like factors in the plant kingdom. Gene. 2013. 526(1): 30-38

Knutson BA, Hahn S. Yeast Rrn7 and human TAF1B are TFIIB-related RNA polymerase I general transcription factors. Science. 2011. 33(6049): 1637-40

Knutson BA, Hahn S. Domains of Tra1 important for activator recruitment and transcription coactivator functions of SAGA and NuA4 complexes. Mol Cell Biol. 2011. 31(4): 818-831

Knutson BA. Insights into the domain and repeat architecture of target of rapamycin. J Struct Biol. 2010. 170(2): 354-63