Department of Functional Genomics

Department of Functional Genomics

Alzheimer’s disease (AD) is a progressive neurodegenerative disorder and the leading cause of senile dementia. There is an urgent need to develop fundamental therapeutic drug for AD. Two physiological anti-Alzheimer factors, BRI2 and BRI3, have been reported to negatively regulate amyloid β (Aβ) production by binding to amyloid precursor protein (APP) and inhibiting its processing by secretases. Moreover, both proteins are also capable of increasing degradation, and decreasing aggregation of Aβ and islet amyloid polypeptide (IAPP), which is known to be related to the pathogenesis of AD in diabetes [Fig. 1]. Of note, two mutations in the BRI2 gene have been identified as the cause of autosomal dominant Familial British and Danish dementias (FBD and FDD, respectively), which share pathological and clinical similarities with AD.

BRI2 gene mutations render mutant BRI2 protein unstable, such that it is mostly degraded. Consequently, proteolytic processing of APP is deregulated, and production of Aβ is increased. In addition, it has been reported recently that the amounts of BRI2 associated with APP are decreased in the hippocampus of AD patients. Based on these findings, it has been suggested that reduced BRI2 functionality may play an important role not only in FBD and FDD development but also in AD pathogenesis.

Recently, we have discovered that NRBP1 homodimerizes and assembles into a Cul2- and Cul4A-containing heterodimeric Cullin-RING ubiquitin ligase (NRBP1-E3) that efficiently targets both BRI2 and BRI3 for ubiquitination in the presence of TSC22D3 and TSC22D4. NRBP1 knockdown in neuronal cells resulted in an increase in the abundance of BRI2 and BRI3, and significantly reduced Aβ production (Yasukawa T., Aso T. et al. Cell Rep. 2020). Thus, disrupting interactions between NRBP1 and its substrates BRI2 and BRI3 may provide a useful therapeutic strategy for AD [Fig. 1].

We are now searching for compounds that specifically inhibit the interaction between NRBP1 and BRI2 through high-throughput screening (HTS) from compound libraries. We will then conduct lead compound generation and optimization based on hit compounds to identify small molecule drug candidates. Finally, we will evaluate the efficacy of obtained compound on brain Aβ burden and memory function using AD model mouse. These studies should provide a solid foundation for the future development of novel AD drug that would inhibit the ubiquitination of BRI2 and BRI3, thereby collectively enhance their multiple anti-Alzheimer roles [Fig. 1, Table 1].

Yasukawa T., Tsutsui A., Tomomori-Sato C., Sato S., Saraf A., Washburn M.P., Florens L., Terada T., Shimizu K., Conaway R.C., Conaway J.W., and Aso T. NRBP1-containing CRL2/CRL4A regulates amyloid β production by targeting BRI2 and BRI3 for degradation.
Cell Rep. 30, 3478-3491, 2020.

Weems J.C., Slaughter B.D., Unruh J.R., Boeing S., Hall S.M., McLaird M.B., Yasukawa T., Aso T., Svejstrup J.Q., Conaway J.W., and Conaway,R.C.　 Cockayne syndrome B protein regulates recruitment of the Elongin A ubiquitin ligase to sites of DNA damage.
J. Biol. Chem. 292, 6431-6437, 2017.

Nakazawa H., Chang K., Shinozaki S., Yasukawa T., Ishimaru K., Yasuhara S., Yu Y.M., Martyn J.A., Tompkins R.G., Shimokado K., Kaneki M. iNOS as a driver of inflammation and apoptosis in mouse skeletal muscle after burn injury: Possible involvement of Sirt1 S-Nitrosylation-mediated acetylation of p65 NF-κB and p53.
PLoS One 12, e0170391, 2017.

Weems J.C., Slaughter B.D., Unruh J.R., Hall S.M., McLaird M.B., Gilmore J.M., Washburn M.P., Florens L., Yasukawa, T., Aso, T., Conaway, J.W., and Conaway, R.C.
Assembly of the Elongin A ubiquitin ligase is regulated by genotoxic and other stresses.
J. Biol. Chem. 290, 15030-15041, 2015.

Kawauchi, J., Inoue, M., Fukuda, M., Uchida, Y., Yasukawa, T., Conaway, R.C., Conaway, J.W., Aso, T., and Kitajima, S. Transcriptional properties of mammalian Elongin A and its role in stress response.
J. Biol. Chem. 288, 24302-24315, 2013.

Yasukawa, T., Bhatt, S., Takeuchi, T., Kawauchi, J., Takahashi, H., Tsutsui, A., Muraoka, T., Inoue, M., Tsuda, M., Kitajima, S., Conaway, R.C., Conaway, J.W., Trainor, P.A., and Aso, T. Transcriptional elongation factor Elongin A regulates retinoic acid-induced gene expression during neuronal differentiation.
Cell Rep. 2, 1129-1136, 2012.

Yamada, K., Tamamori-Adachi, M., Goto, I., Iizuka, M., Yasukawa, T., Aso, T., Okazaki, T., and Kitajima, S. Degradation of p21Cip1 through APC/CCdc20 mediated ubiquitylation is inhibited by cyclin dependent kinase 2 in cardiomyocytes.
J. Biol. Chem. 286, 44057-44066, 2011.

Yasukawa, T., Kamura, T., Kitajima, S., Conaway, R.C., Conaway, J.W., and Aso, T.
Mammalian Elongin A complex mediates DNA-damage-induced ubiquitylation and degradation of Rpb1.
EMBO J. 27, 3256-3266, 2008. (Faculty of 1000 Biology)

Miyata, K.¶, Yasukawa, T.¶, Fukuda, M., Takeuchi, T., Yamazaki, K., Sakumi, K., Tamamori-Adachi, M., Ohnishi, Y., Ohtsuki, Y., Nakabeppu, Y., Kitajima, S., Onishi, S., and Aso, T.
Induction of apoptosis and cellular senescence in mice lacking transcription elongation factor, Elongin A.
Cell Death Differ. 14, 716-726, 2007. (¶ equal contribution)

Yasukawa, T., Sugimura, K., Fukuda, M., Yamazaki, K., Kitajima, S., Okumura, K., and Aso, T.
Functional characterization of a mammalian transcription factor, Elongin A.
Biochem. Biophys. Res. Commun. 352, 237-243, 2007.

Sugita, H., Fujimoto, M., Yasukawa, T., Shimizu, N., Sugita, M., Yasuhara, S., Martytn, J.A. and Kaneki, M. Inducible nitric-oxide synthase and NO donor induce insulin receptor substrate-1 degradation in skeletal muscle cells.
J. Biol. Chem. 280, 14203-14211, 2005.

Yasukawa, T., Tokunaga, E., Ota, H., Sugita, H., Martyn, J.A. and Kaneki, M.
S-Nitrosylation-dependent inactivation of Akt/PKB in insulin resistance.
J. Biol. Chem. 280, 7511-7518, 2005.

Yamazaki, K., Aso, T., Ohnishi, Y., Ohno, M., Tamura, K., Shuin, T., Kitajima, S., and Nakabeppu, Y. Mammalian Elongin A is not essential for cell viability but is required for proper cell-cycle progression with limited alteration of gene expression.
J. Biol. Chem. 278, 13585-13589, 2003.

Yamazaki, K., Guo, L., Sugahara, K., Zhang, C., Enzan, H., Nakabeppu, Y., Kitajima, S., and Aso, T. Identification and biochemical characterization of a novel transcription elongation factor, Elongin A3.
J. Biol. Chem. 277, 26444-26451, 2002.

Zhang, C., Yong, C., Adachi, M.T., Oshiro, S., Aso, T., Kaufman, R.J., and Kitajima, S.
Homocysteine induces programmed cell death in human vascular endothelial cells through activation of the unfolded protein response.
J. Biol. Chem. 276, 35867-35874, 2001.

Matsuda, S., Yasukawa, T., Homma, Y., Shao, Z., Ito, Y., Niikura, T., Hiraki, T., Hirai, S., Ohno, S., Kita, Y., Kawasumi, M., Kouyama, K., Yamamoto, T., Kyriakis, J.M., Nishimoto, I. 　c-Jun N-terminal kinase (JNK)-interacting protein-1β/islet-brain-1 scaffolds Alzheimer's amyloid precursor protein with JNK. J.Neurosci. 21, 6597-6607, 2001.

Hashimoto, Y., Niikura, T., Tajima, H., Yasukawa, T., Sudo, H., Ito, Y., Kita, Y., Kawasumi, M., Kouyama, K., Doyu, M., Sobue, G., Koide, T., Tsuji, S., Lang, J., Kurokawa, K., Nishimoto, I. A rescue factor abolishing neuronal cell death by a wide spectrum of familial Alzheimer's disease genes and Aβ.
Proc. Natl. Acad. Sci. U. S. A. 98, 6336-6341, 2001.

Sudo, H., Hashimoto, Y., Niikura, T., Shao, Z., Yasukawa, T., Ito, Y., Yamada, M., Hata, M., Hiraki, T., Kawasumi, M., Kouyama, K., and Nishimoto, I. Secreted Aβ does not mediate neurotoxicity by antibody-stimulated amyloid precursor protein.
Biochem. Biophys. Res. Commun. 282, 548-556, 2001.

Niikura, T., Hashimoto, Y., Okamoto, T., Abe, Y., Yasukawa, T., Kawasumi, M., Hiraki, T., Kita, Y., Terashita, K., Kouyama, K. and Nishimoto, I. GF-I protects cells from apoptosis by Alzheimer's V642I mutant APP through IGF-I receptor in an IGF binding protein-sensitive manner.
J. Neurosci. 21, 1902-1910, 2001.

Kokura, K., Kaul, S.C., Wadhwa, R., Nomura, T., Khan, M.M., Shinagawa, T., Yasukawa, T., Colmanares, C., Ishii, S. The Ski protein family is required for MeCP2-mediated transcriptional repression.
J. Biol. Chem. 276, 34115-34121, 2001.

Cai, Y., Zhang, C., Nawa, T., Aso, T., Tanaka, M., Oshiro, S., Ichijo, H., and Kitajima, S. Homocysteine-responsive ATF3 gene expression in human vascular endothelial cells: signal transduction pathway and promoter responsive element.
Blood 96, 2140-2148, 2000.

Aso, T., Yamazaki, K., Amimoto, K., Kuroiwa, A., Higashi, H., Matsuda, Y., Kitajima, S., and Hatakeyama, M. Identification and characterization of Elongin A2, a new member of an Elongin family of transcription elongation factors, specifically expressed in the testis.
J. Biol. Chem. 275, 6546-6552, 2000.

Aso, T., Yamazaki, K., Aigaki, T., and Kitajima, S.
Drosophila von Hippel-Lindau tumor suppressor complex possesses E3 ubiquitin ligase activity.
Biochem. Biophys. Res. Commun. 276, 355-361, 2000.

Sudo, H. ¶, Jiang, H. ¶, Yasukawa, T. ¶, Hashimoto, Y., Niikura, T., Kawasumi, M., Matsuda, S., Takeuchi, Y., Aiso, S., Matsuoka, M., Murayama, Y. and Nishimoto, I.
Antibody regulated neurotoxic function of cell surface β-amyloid precursor protein.
Mol. Cell. Neurosci. 16, 708-723, 2000. (¶ equal contribution)

Ohh, M., Takagi, Y., Aso, T., Stebbins, C.E., Pavletich, N.P., Zbar, B., Conaway, R.C., Conaway, J.W., and Kaelin, W.G. Synthetic peptides define critical contacts between elongin C, elongin B, and the von Hippel-Lindau tumor suppressor protein.
J. Clin. Invest. 104, 1583-1591, 1999.

Shinobu, N., Maeda, T., Aso, T., Ito, T., Kondo, T., Koike, K., and Hatakeyama, M. Physical interaction and functional antagonism between the RNA polymerase II elongation factor ELL and p53.
J. Biol. Chem. 274, 17003-17010, 1999.

Tokitou, F., Nomura, T., Khan, M.M., Kanl, S.C., Wadhwa, K., Yasukawa, T., Kohno, I. and Ishii, S. Viral ski inhibits retinoblastoma protein (Rb)-mediated transcriptional repression in a dominant negative fashion.
J. Biol. Chem. 274, 4485-4488, 1999.

Aso, T., and Conrad, M.N. Molecular cloning of DNAs encoding the regulatory subunits of Elongin from Saccharomyces cerevisiae and Drosophila melanogaster.
Biochem. Biophys. Res. Commun. 241, 334-340, 1997.

Pan, G., Aso, T., and Greenblatt, J.
Interaction of elongation factors TFIIS and elongin A with a human RNA polymerase II holoenzyme capable of promoter-specific initiation and responsive to transcriptional activators.
J. Biol. Chem. 272, 24563-24571, 1997.

Aso, T., Haque, D., Barstead, R.J., Conaway, R.C., and Conaway, J.W. The inducible elongin A elongation activation domain: structure, function and interaction with the elongin BC complex.
EMBO J. 15, 5557-5566, 1996.

Aso, T., Shilatifard, A., Conaway, J.W., and Conaway, R.C.
Transcription syndromes and the role of RNA polymerase II general transcription factors in human disease.
J. Clin. Invest. 97, 1561-1569, 1996.

Dai, P., Akimaru, H., Tanaka, T., Hou, D-X., Yasukawa, T., Kanei-Ishi, C., Takahashi, T. and Ishii, S.
CBP as a transcriptional coactivator of c-Myb.
Genes Dev. 1, 528-540, 1996.

Aso, T., Lane, W.S., Conaway, J.W., and Conaway, R.C.
Elongin (SIII) : A multisubunit regulator of elongation by RNA polymerase II.
Science 269, 1439-1443, 1995.

Duan, D.R., Pause, A., Burgess, W.H., Aso, T., Chen, D.Y.T., Garrett, K.P., Conaway, R.C., Conaway, J.W., Linehan, W.M., and Klausner, R.D.
Inhibition of transcription elongation by the von Hippel-Lindau tumor suppressor protein.
Science 269, 1402-1406, 1995.

Garrett, K.P. ¶, Aso, T. ¶, Bradsher, J.N., Foundling, S.I., Lane, W.S., Conaway, R.C., and Conaway, J.W. Positive regulation of general transcription factor SIII by a tailed ubiquitin homolog.
Proc. Natl. Acad. Sci. U. S. A. 92, 7172-7176, 1995. (¶ equal contribution)

Aso, T., Conaway, J.W., and Conaway, R.C. The RNA polymerase II elongation complex.
FASEB J. 9, 1419-1428, 1995.

Mizuguchi, G., Kanei-Ishii, C., Takahashi, T., Yasukawa, T., Nagase, T., Horikoshi, M., Yamamoto, T. and Ishii, S. c-Myb repression of c-erbB-2 transcription by direct binding to the c-erbB-2 promoter.
J. Biol. Chem. 270, 9384-9389, 1995.

Yasukawa, T., Kanei-Ishii, C., Maekawa, T., Fujimoto, J., Yamamoto, T. and Ishii, S.
Increase of solubility of foreign proteins in Eschericha coli by coproduction of the bacterial thioredoxin.
J. Biol. Chem. 270, 25328-25331, 1995.

Aso, T., Serizawa, H., Conaway, R.C., and Conaway, J.W. A TATA sequence-dependent transcriptional repressor activity associated with mammalian transcription factor IIA.
EMBO J. 13, 435-445, 1994.

Aso, T., Conaway, J.W., and Conaway, R.C.
Role of core promoter structure in assembly of the RNA polymerase II preinitiation complex.
J. Biol. Chem. 269, 26575-26583, 1994.

Tan, S, Aso, T., Conaway, R.C., and Conaway, J.W. Roles of both the RAP30 and RAP74 subunits of transcription factor IIF in transcription initiation and elongation by RNA polymerase II.
J. Biol. Chem. 269, 25684-25691, 1994.

Kanei-Ishii, C., Yasukawa, T., Morimoto, R.I. and Ishii, S. c-Myb-induced trans activation mediated by heat shock elements without sequence-specific DNA binding of c-Myb.
J. Biol. Chem. 269, 15768-15775, 1994.

Yonaha, M., Aso, T., Kobayashi, Y., Vasavada, H., Yasukochi, Y., Weissman, S.M., and Kitajima, S. Domain structure of a human general transcription initiation factor, TFIIF.
Nucleic Acid Res. 21, 273-279, 1993.

Sarai, A., Uedaira, H., Morii, H., Yasukawa, T., Ogata, K., Nishimura, Y. and Ishii, S.
Thermal stability of the DNA-binding domain of the Myb oncoprotein.
Biochemistry 39, 7759-7764, 1993.

Tanikawa, J., Yasukawa, T., Enari, M., Ogata, K., Nishimura, Y., Ishii, S. and Sarai, A.
Recognition of specific DNA sequences by the c-myb protooncogene product: Role of three repeat units in the DNA-binding domain.
Proc. Natl. Acad. Sci. U. S. A. 90, 9320-9324, 1993.

Aso, T., Vasavada, H., Kawaguchi, T., Germino, F.J., Ganguly, S., Kitajima, S., Weissman, S.M., and Yasukochi, Y. Characterization of cDNA for the large subunit of the transcription initiation factor TFIIF.
Nature 355, 461-464, 1992.