Intrinsic bioenergetic adaptations compensate for reduced mitochondrial content in HER2-driven mammary tumors

Reviewer #1 (Public review):

Summary:

In this manuscript, Frangos et al. used a transcriptomic and proteomic approach to characterise changes in HER2-driven mammary tumours compared to healthy mammary tissue in mice. They observed that mitochondrial genes, including OXPHOS regulators, were among the most down-regulated genes and proteins in their datasets. Surprisingly, these were associated with higher mitochondrial respiration, in response to a variety of carbon sources. In addition, there seems to be a reduction in mitochondrial fusion and an increase in fission in tumours compared to healthy tissues.

Strengths:

The data are clearly presented and described.

The author reported very similar trends in proteomic and transcriptomic data. Such approaches are essential to have a better understanding of the changes in cancer cell metabolism associated with tumourigenesis

Weaknesses:

This study, despite being a useful resource (assuming all the data will be publicly available and not only upon request) is mainly descriptive and correlative and lacks mechanistic links.

It would be important to determine the cellular composition of the tumour and healthy tissue used. Do the changes described here apply to cancer cells only or do other cell types contribute to this?

Are the changes in metabolic gene expression a consequence of HER2 signalling activation? Ex-vivo experiments could be performed to perturb this pathway and determine cause-effects.

The data of fission/fusion seem quite preliminary and the gene/protein expression changes are not so clear cut to be a convincing explanation that this is the main reason for the increased mitochondria respiration in tumours.

Reviewer #2 (Public review):

Frangos et al present a set of studies aiming to determine mechanisms underlying initiation and tumour progression. Overall, this work provides some useful insights into the involvement of mitochondrial dysfunction during the cellular transformation process. This body of work could be improved in several possible directions to establish more mechanistic connections.

(1) The interesting point of the paper: the contrast between suppressed ETC components and activated OXPHOS function is perplexing and should be resolved. It is still unclear if activated mitochondrial function triggers gene down-regulation vs compensatory functional changes (as the title suggests). Have the authors considered reversing the HER2-derived signals e.g. with PI3K-AKT-MTOR or ERK inhibitors to potentially separate the expression vs. functional phenotypes? The root of the OXPHOS component down-regulation should also be traced further, e.g. by probing into levels of core mitochondrial biogenesis factors. Are transcript levels of factors encoded by mtDNA also decreased?

(2) The second interesting aspect of this study is the implication of mitochondrial activation in tumours, despite the downregulation of expression signatures, suggestive of a positive role for mitochondria in this tumour model. To address if this is correlative or causal, have the authors considered testing an OXPHOS inhibitor for suppression of tumorigenesis?

(3) A number of issues concerning animal/ tumour variability and further pathway dissection could be explored with in vitro approaches. Have the authors considered deriving tumour-derived cell cultures, which could enable further confirmations, mechanistic drug studies and additional imaging approaches? Culture systems would allow alternative assessment of mitochondrial function such as Seahorse or flow cytometry (mitochondrial potential and ROS levels).

(4) The study could be greatly improved with further confirmatory studies, eg immunoblotting for mitochondrial components with parallel blots for phospho-signalling in the same samples. It would be interesting if trends could be maintained in tumour-derived cell cultures. It is notable that OXPHOS protein/transcript changes are more consistent (Figure 5, Supplementary Figure 4) than mitochondrial dynamics /mitophagy factors (Figure 8). Core regulatory factors in these pathways should be confirmed by conventional immunoblotting.