A/Prof Ruth Kluck, Lab Head | WEHI Researcher Profile

Q: Understanding mitochondrial pore formation during apoptotic cell death

A key event in apoptotic cell death is the oligomerisation of the BAX and BAK proteins to form pores in mitochondria, although how they form pores is still unclear.We recently found that cells lacking the putative trafficking protein PACS1 are resistant to apoptosis due to unusual complexes of BAX and BAK (Brasacchio et al, Cell Death Differ, 2017).We are thus characterising the unusual BAX and BAK complexes in PACS1-knockdown cells to understand this new means of resistance.

Q: Elucidating how homodimers of BAX and of BAK form the apoptotic pore

As the formation of BAX and BAK homo-oligomers strongly correlates with their ability to perforate mitochondria, defining how BAX and BAK dimers self-associate and interact with the membrane will reveal how they trigger apoptosis.Our data indicate that dimers do not interact by distinct protein-protein interface, but form disordered clusters to generate pores (Uren et al, eLife, 2017; Uren et al, Philos Trans R Soc Lond B Biol Sci, 2017).A range of biochemical approaches will examine further how the outer membrane is involved in oligomerisation of dimers.

Q: Determining how MCL-1 contributes to resistance during anti-cancer treatment

Inhibition of apoptosis by prosurvival BCL-2 proteins contributes to oncogenesis and to resistance to cancer treatments. In particular, MCL-1 can cause resistance by sequestering activated BAK.We aim to better understand when and how MCL-1 and BAK interact in different cancer cells following treatment, and so identify ways of circumventing this resistance.

Q: Delivering antibodies into cells to trigger apoptotic cell death

We found that an antibody to the BAK protein can trigger its activation leading to mitochondrial pore formation and cell death (Iyer et al, Nat Commun 2016 7:11734).To investigate if this antibody can be developed as a novel anti-cancer agent, this project will combine the anti-BAK antibody with others that can be taken up by cancer cells, and test for induction of cell death.

About

Our research program uses biochemical, cell biological and structural approaches to examine how the BCL-2 protein family regulates the mitochondrial pathway of apoptosis.

We are particularly interested in how two pore-forming BCL-2 family members, BAX and BAK, undergo conformation change and generate pores in the mitochondrial outer membrane. The goal of our research is to identify new means of specifically regulating apoptotic cell death in cancer and other diseases.

We have found that following major conformation changes, both BAX and BAK form unusual symmetric homodimers that act as the basic unit of the apoptotic pore, and are using different strategies to determine how dimers form clusters that disrupt the membrane.

We also discovered that BAK can be activated by antibodies, and are testing whether these antibodies can be delivered into cancer cells to directly trigger BAK-mediated apoptosis.

Education

Grants and funding

Publications

Selected publications from A/Prof Ruth Kluck

Cummings MC, Vaux DL, Strasser A, Kluck R. Correction: John F. R. Kerr (1934–2024). Cell Death & Differentiation. 2024;31(10):10.1038/s41418-024-01373-8

Cummings MC, Vaux DL, Strasser A, Kluck R. John F. R. Kerr (1934–2024). Cell Death & Differentiation. 2024;31(8):10.1038/s41418-024-01338-x

Subas Satish HP, Iyer S, Shi MX, Wong AW, Fischer KC, Wardak AZ, Lio D, Brouwer JM, Uren RT, Czabotar PE, Miller MS, Kluck RM. A novel inhibitory BAK antibody enables assessment of non-activated BAK in cancer cells. Cell Death & Differentiation. 2024;31(6):10.1038/s41418-024-01289-3

Birkinshaw RW, Iyer S, Lio D, Luo C, Brouwer J, Miller MS, Robin A, Uren RT, Dewson G, Kluck RM, Colman PM, Czabotar PE. The crystal structure of detergent-activated BAK dimers provides insights into BAK activation steps and how they stabilise membrane pores. Acta Crystallographica Section A: Foundations and advances. 2023;79(a2):10.1107/s205327332308912x

Miller MS, Subas-Satish H, Iyer S, Wardak AZ, Lio D, Shi MX, Wong A, Uren R, Brouwer JM, Czabotar PE, Kluck RM. Structure of the BAK- inhibiting antibody 14G6 bound to BAK identifies core–latch separation and α1-dissociation as essential events in BAK activation. Acta Crystallographica Section A: Foundations and advances. 2023;79(a2):10.1107/s2053273323086709

Vitale I, Pietrocola F, Guilbaud E, Aaronson SA, Abrams JM, Adam D, Agostini M, Agostinis P, Alnemri ES, Altucci L, Amelio I, Andrews DW, Aqeilan RI, Arama E, Baehrecke EH, Balachandran S, Bano D, Barlev NA, Bartek J, Bazan NG, Becker C, Bernassola F, Bertrand MJM, Bianchi ME, Blagosklonny MV, Blander JM, Blandino G, Blomgren K, Borner C, Bortner CD, Bove P, Boya P, Brenner C, Broz P, Brunner T, Damgaard RB, Calin GA, Campanella M, Candi E, Carbone M, Carmona-Gutierrez D, Cecconi F, Chan FK-M, Chen G-Q, Chen Q, Chen YH, Cheng EH, Chipuk JE, Cidlowski JA, Ciechanover A, Ciliberto G, Conrad M, Cubillos-Ruiz JR, Czabotar PE, D’Angiolella V, Daugaard M, Dawson TM, Dawson VL, De Maria R, De Strooper B, Debatin K-M, Deberardinis RJ, Degterev A, Del Sal G, Deshmukh M, Di Virgilio F, Diederich M, Dixon SJ, Dynlacht BD, El-Deiry WS, Elrod JW, Engeland K, Fimia GM, Galassi C, Ganini C, Garcia-Saez AJ, Garg AD, Garrido C, Gavathiotis E, Gerlic M, Ghosh S, Green DR, Greene LA, Gronemeyer H, Häcker G, Hajnóczky G, Hardwick JM, Haupt Y, He S, Heery DM, Hengartner MO, Hetz C, Hildeman DA, Ichijo H, Inoue S, Jäättelä M, Janic A, Joseph B, Jost PJ, Kanneganti T-D, Karin M, Kashkar H, Kaufmann T, Kelly GL, Kepp O, Kimchi A, Kitsis RN, Klionsky DJ, Kluck R, Krysko DV, Kulms D, Kumar S, Lavandero S, Lavrik IN, Lemasters JJ, Liccardi G, Linkermann A, Lipton SA, Lockshin RA, López-Otín C, Luedde T, MacFarlane M, Madeo F, Malorni W, Manic G, Mantovani R, Marchi S, Marine J-C, Martin SJ, Martinou J-C, Mastroberardino PG, Medema JP, Mehlen P, Meier P, Melino G, Melino S, Miao EA, Moll UM, Muñoz-Pinedo C, Murphy DJ, Niklison-Chirou MV, Novelli F, Núñez G, Oberst A, Ofengeim D, Opferman JT, Oren M, Pagano M, Panaretakis T, Pasparakis M, Penninger JM, Pentimalli F, Pereira DM, Pervaiz S, Peter ME, Pinton P, Porta G, Prehn JHM, Puthalakath H, Rabinovich GA, Rajalingam K, Ravichandran KS, Rehm M, Ricci J-E, Rizzuto R, Robinson N, Rodrigues CMP, Rotblat B, Rothlin CV, Rubinsztein DC, Rudel T, Rufini A, Ryan KM, Sarosiek KA, Sawa A, Sayan E, Schroder K, Scorrano L, Sesti F, Shao F, Shi Y, Sica GS, Silke J, Simon H-U, Sistigu A, Stephanou A, Stockwell BR, Strapazzon F, Strasser A, Sun L, Sun E, Sun Q, Szabadkai G, Tait SWG, Tang D, Tavernarakis N, Troy CM, Turk B, Urbano N, Vandenabeele P, Vanden Berghe T, Vander Heiden MG, Vanderluit JL, Verkhratsky A, Villunger A, von Karstedt S, Voss AK, Vousden KH, Vucic D, Vuri D, Wagner EF, Walczak H, Wallach D, Wang R, Wang Y, Weber A, Wood W, Yamazaki T, Yang H-T, Zakeri Z, Zawacka-Pankau JE, Zhang L, Zhang H, Zhivotovsky B, Zhou W, Piacentini M, Kroemer G, Galluzzi L. Apoptotic cell death in disease—Current understanding of the NCCD 2023. Cell Death & Differentiation. 2023;30(5):10.1038/s41418-023-01153-w

Diepstraten ST, Young S, La Marca JE, Wang Z, Kluck RM, Strasser A, Kelly GL. Lymphoma cells lacking pro-apoptotic BAX are highly resistant to BH3-mimetics targeting pro-survival MCL-1 but retain sensitivity to conventional DNA-damaging drugs. Cell Death & Differentiation. 2023;30(4):10.1038/s41418-023-01117-0

Satish HPS, Zeglinski K, Uren RT, Nutt SL, Ritchie ME, Gouil Q, Kluck RM. NAb-seq: an accurate, rapid, and cost-effective method for antibody long-read sequencing in hybridoma cell lines and single B cells. mAbs. 2022;14(1):10.1080/19420862.2022.2106621

Robin AY, Miller MS, Iyer S, Shi MX, Wardak AZ, Lio D, Smith NA, Smith BJ, Birkinshaw RW, Czabotar PE, Kluck RM, Colman PM. Structure of the BAK-activating antibody 7D10 bound to BAK reveals an unexpected role for the α1-α2 loop in BAK activation. Cell Death & Differentiation. 2022;29(9):10.1038/s41418-022-00961-w

Ke FS, Holloway S, Uren RT, Wong AW, Little MH, Kluck RM, Voss AK, Strasser A. The BCL‐2 family member BID plays a role during embryonic development in addition to its BH3‐only protein function by acting in parallel to BAX, BAK and BOK. The EMBO Journal. 2022;41(15):10.15252/embj.2021110300