The class II myosin MYH4 safeguards genome integrity and suppresses tumor progression
Jayashree Thatte, Ana Moisés da Silva, Judit Börcsök, Thorkell Gudjónsson, Jan Benada, Xin Li, Muthiah Bose, Hanneke van der Gulden, Ji-Ying Song, Renée Menezes, Elena Martín-Doncel, Luis Toledo, Valdemaras Petrosius, Cord Brakebusch, Jos Jonkers, Finn Cilius Nielsen, Maria Rossing, Claus S. Sørensen
Jayashree Thatte, Ana Moisés da Silva, Judit Börcsök, Thorkell Gudjónsson, Jan Benada, Xin Li, Muthiah Bose, Hanneke van der Gulden, Ji-Ying Song, Renée Menezes, Elena Martín-Doncel, Luis Toledo, Valdemaras Petrosius, Cord Brakebusch, Jos Jonkers, Finn Cilius Nielsen, Maria Rossing, Claus S. Sørensen
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Research Article Cell biology Clinical Research Oncology

The class II myosin MYH4 safeguards genome integrity and suppresses tumor progression

Abstract

Loss-of-function mutations in genome maintenance genes fuel tumorigenesis through increased genomic instability. A subset of these tumor suppressors are challenging to identify due to context dependency, including functional interactions with other genes and pathways. Here, we searched for potential causal genes that impact tumor development and/or progression in breast cancer through functional-genetic screening of candidate genes. MYH4, encoding a class II myosin, emerged as a top hit impacting genomic stability. We show that MYH4 suppresses DNA replication stress by promoting replication licensing and replication fork progression. Moreover, we observed a strong synergistic relationship among class II myosins in suppressing replication-associated DNA damage. Genomic analysis of Pan-Cancer Analysis of Whole Genomes project breast cancer samples revealed frequent concomitant loss of TP53 with MYH4 and class II myosins on chromosome 17p. Notably, Myh4 disruption accelerated mouse mammary tumorigenesis in a Trp53-deficient background. In conclusion, our results suggest an unanticipated function of MYH4 in p53-mediated tumor suppression that can explain their combined loss in breast cancer.

Authors

Jayashree Thatte, Ana Moisés da Silva, Judit Börcsök, Thorkell Gudjónsson, Jan Benada, Xin Li, Muthiah Bose, Hanneke van der Gulden, Ji-Ying Song, Renée Menezes, Elena Martín-Doncel, Luis Toledo, Valdemaras Petrosius, Cord Brakebusch, Jos Jonkers, Finn Cilius Nielsen, Maria Rossing, Claus S. Sørensen

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Figure 1

MYH4 supports genomic integrity.

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MYH4 supports genomic integrity. (A and B) Scatter plots depicting genes...
(A and B) Scatter plots depicting genes that scored with 2 or more siRNAs for cell fitness phenotype in U2OS and MCF10A breast cell lines, respectively. MYH4 is highlighted in red. The y axis shows the P values calculated from the mean of 3 replicates; the x axis shows scores for relative cell number, normalized to siUNC = 1. (C) U2OS cells transfected with siUNC or siMYH4 (no. 3) for 72 hours and subjected to immunofluorescence analysis to assess γH2AX levels. Representative confocal images are shown; DAPI was used to stain nuclei. Scale bar: 20 μm. (D) QIBC plot of U2OS cells transfected with indicated siRNAs and stained for γH2AX. Plot shows mean intensity of γH2AX per nucleus on the y axis and total DNA content on the x axis. Color legend: green, minimum; yellow, average; red, maximum. (E) A bar chart showing relative γH2AX mean intensity of U2OS cells transfected with siMYH4 (no. 3). Data points show 7 biological replicates. Data presented as mean ± SD. (F) Representative images of alkaline comet assay from 3 biological replicates. Ionizing radiation–treated (IR-treated) cells were used as a positive control. Scale bar: 20 μm. (G) Quantification of comet tail moment from 3 biological replicates. Error bars indicate ±SEM. (H) Confocal images showing micronuclei formation (red arrows) after MYH4 knockdown; DAPI was used to stain nuclei. Scale bar: 20 μm. (I) Quantification of cells harboring micronuclei; relative values from 5 biological replicates are shown. Data presented as mean ± SD. *P < 0.05; **P < 0.001; ***P < 0.001 by 2-tailed, paired t test.