p66ShcA functions as a contextual promoter of breast cancer metastasis

Abstract

Background: The p66ShcA redox protein is the longest isoform of the Shc1 gene and is variably expressed in breast cancers. In response to a variety of stress stimuli, p66ShcA becomes phosphorylated on serine 36, which allows it to translocate from the cytoplasm to the mitochondria where it stimulates the formation of reactive oxygen species (ROS). Conflicting studies suggest both pro-and anti-tumorigenic functions for p66ShcA, which prompted us to examine the contribution of tumor cell-intrinsic functions of p66ShcA during breast cancer metastasis. Methods: We tested whether p66ShcA impacts the lung-metastatic ability of breast cancer cells. Breast cancer cells characteristic of the ErbB2+/luminal (NIC) or basal (4T1) subtypes were engineered to overexpress p66ShcA. In addition, lung-metastatic 4T1 variants (4T1-537) were engineered to lack endogenous p66ShcA via Crispr/Cas9 genomic editing. p66ShcA null cells were then reconstituted with wild-type p66ShcA or a mutant (S36A) that cannot translocate to the mitochondria, thereby lacking the ability to stimulate mitochondrial-dependent ROS production. These cells were tested for their ability to form spontaneous metastases from the primary site or seed and colonize the lung in experimental (tail vein) metastasis assays. These cells were further characterized with respect to their migration rates, focal adhesion dynamics, and resistance to anoikis in vitro. Finally, their ability to survive in circulation and seed the lungs of mice was assessed in vivo. Results: We show that p66ShcA increases the lung-metastatic potential of breast cancer cells by augmenting their ability to navigate each stage of the metastatic cascade. A non-phosphorylatable p66ShcA-S36A mutant, which cannot translocate to the mitochondria, still potentiated breast cancer cell migration, lung colonization, and growth of secondary lung metastases. However, breast cancer cell survival in the circulation uniquely required an intact p66ShcA S36 phosphorylation site. Conclusion: This study provides the first evidence that both mitochondrial and non-mitochondrial p66ShcA pools collaborate in breast cancer cells to promote their maximal metastatic fitness.

The SHCA adapter protein cooperates with lipoma-preferred partner in the regulation of adhesion dynamics and invadopodia formation

“…Accordingly, lung metastatic burden is significantly reduced when SHCA signaling is disrupted (31,32). This data is consistent with findings in triple negative breast cancer cells where p66SHCA is required for efficient cell migration and lung metastasis (35). Loss of p66SHCA expression results in the formation of large, elongated adhesions that exhibit slower assembly and disassembly rates whereas exogenous expression of p66SHCA induces an EMT in ErbB2 + luminal breast cancers (34,35).…”

“…All breast cancer cells express p46/52 SHCA isoforms, while p66SHCA is more highly expressed in breast cancers with mesenchymal features (34). Recently, p66SHCA has been shown to be a context-dependent promoter of breast cancer metastasis (35). Loss of p66SHCA expression results in slower adhesion dynamics, reduced cell migration rates, and diminished lung metastasis (35).…”

“…Recently, p66SHCA has been shown to be a context-dependent promoter of breast cancer metastasis (35). Loss of p66SHCA expression results in slower adhesion dynamics, reduced cell migration rates, and diminished lung metastasis (35). SHCA harbors an Nterminal phosphotyrosine binding domain (PTB), a central collagen homology domain (CH1) with three key tyrosine residues (Y239/Y240/Y313) and a C-terminal SH2 domain (33).…”

“…To verify the results of our semiautomatic analysis, a custom algorithm was created in MATLAB (35,115). Here, a spline curve was first fitted to each intensity trace to identify segments of assembly and disassembly.…”

Photodynamic therapy regulates fate of cancer stem cells through reactive oxygen species

“…When it undergoes oxidative stress, p66Shc is phosphorylated at Ser36 and then is translocated to mitochondria and/or MAMs. It is involved in ROS generation and apoptosis-related signaling pathways[ 160 , 161 ].…”