DNA damage and tissue repair: What we can learn from planaria.

Faithful renewal of aging and damaged tissues is central to organismal lifespan. Stem cells (SCs) generate the cellular progeny that replenish adult tissues across the body but this task becomes increasingly compromised over time. The age related decline in SC-mediated tissue maintenance is a multifactorial event that commonly affects genome integrity. The presence of DNA damage in SCs that are under continuous demand to divide poses a great risk for age-related disorders such as cancer. However, performing analysis of SCs with genomic instability and the DNA damage response during tissue renewal present significant challenges. Here we introduce an alternative experimental system based on the planaria flatworm Schmidtea mediterranea to address at the organismal level studies intersecting SC-mediated tissue renewal in the presence of genomic instability. Planaria have abundant SCs (neoblasts) that maintain high rates of cellular turnover and a variety of molecular tools have been developed to induce DNA damage and dissect how neoblasts respond to this stressor. S. mediterranea displays high evolutionary conservation of DNA repair mechanisms and signaling pathways regulating adult SCs. We describe genetically induced-DNA damage models and highlight body-wide signals affecting cellular decisions such as survival, proliferation, and death in the presence of genomic instability. We also discuss transcriptomic changes in the DNA damage response during injury repair and propose DNA repair as key component of tissue regeneration. Additional studies using planaria will provide insights about mechanisms regulating survival and growth of cells with DNA damage during tissue renewal and regeneration

SUMOylation controls stem cell proliferation and regional cell death through Hedgehog signaling in planarians.

Mechanisms underlying anteroposterior body axis differences during adult tissue maintenance and regeneration are poorly understood. Here, we identify that post-translational modifications through the SUMO (Small Ubiquitin-like Modifier) machinery are evolutionarily conserved in the Lophotrocozoan Schmidtea mediterranea. Disruption of SUMOylation in adult animals by RNA-interference of the only SUMO E2 conjugating enzyme Ubc9 leads to a systemic increase in DNA damage and a remarkable regional defect characterized by increased cell death and loss of the posterior half of the body. We identified that Ubc9 is mainly expressed in planarian stem cells (neoblasts) but it is also transcribed in differentiated cells including neurons. Regeneration in Ubc9(RNAi) animals is impaired and associated with low neoblast proliferation. We present evidence indicating that Ubc9-induced regional cell death is preceded by alterations in transcription and spatial expression of repressors and activators of the Hedgehog signaling pathway. Our results demonstrate that SUMOylation acts as a regional-specific cue to regulate cell fate during tissue renewal and regeneration

Cancer epithelia-derived mitochondrial DNA is a targetable initiator of a paracrine signaling loop that confers taxane resistance.

Stromal-epithelial interactions dictate cancer progression and therapeutic response. Prostate cancer (PCa) cells were identified to secrete greater concentration of mitochondrial DNA (mtDNA) compared to noncancer epithelia. Based on the recognized coevolution of cancer-associated fibroblasts (CAF) with tumor progression, we tested the role of cancer-derived mtDNA in a mechanism of paracrine signaling. We found that prostatic CAF expressed DEC205, which was not expressed by normal tissue-associated fibroblasts. DEC205 is a transmembrane protein that bound mtDNA and contributed to pattern recognition by Toll-like receptor 9 (TLR9). Complement C3 was the dominant gene targeted by TLR9-induced NF-κB signaling in CAF. The subsequent maturation complement C3 maturation to anaphylatoxin C3a was dependent on PCa epithelial inhibition of catalase in CAF. In a syngeneic tissue recombination model of PCa and associated fibroblast, the antagonism of the C3a receptor and the fibroblastic knockout of TLR9 similarly resulted in immune suppression with a significant reduction in tumor progression, compared to saline-treated tumors associated with wild-type prostatic fibroblasts. Interestingly, docetaxel, a common therapy for advanced PCa, further promoted mtDNA secretion in cultured epithelia, mice, and PCa patients. The antiapoptotic signaling downstream of anaphylatoxin C3a signaling in tumor cells contributed to docetaxel resistance. The inhibition of C3a receptor sensitized PCa epithelia to docetaxel in a synergistic manner. Tumor models of human PCa epithelia with CAF expanded similarly in mice in the presence or absence of docetaxel. The combination therapy of docetaxel and C3 receptor antagonist disrupted the mtDNA/C3a paracrine loop and restored docetaxel sensitivity

Antagonizing Glutamine Bioavailability Promotes Radiation Sensitivity in Prostate Cancer

Nearly half of localized prostate cancer (PCa) patients given radiation therapy develop recurrence. Here, we identified glutamine as a key player in mediating the radio-sensitivity of PCa. Glutamine transporters and glutaminase are upregulated by radiation therapy of PCa cells, but respective inhibitors were ineffective in radio-sensitization. However, targeting glutamine bioavailability by L-asparaginase (L-ASP) led to a significant reduction in clonogenicity when combined with irradiation. L-ASP reduced extracellular asparagine and glutamine, but the sensitization effects were driven through its depletion of glutamine. L-ASP led to G2/M cell cycle checkpoint blockade. As evidence, there was a respective delay in DNA repair associated with RAD51 downregulation and upregulation of CHOP, contributing to radiation-induced cell death. A radio-resistant PCa cell line was developed, was found to bypass radiation-induced mitotic catastrophe, and was sensitive to L-ASP/radiation combination treatment. Previously, PCa-associated fibroblasts were reported as a glutamine source supporting tumor progression. As such, glutamine-free media were not effective in promoting radiation-induced PCa cell death when co-cultured with associated primary fibroblasts. However, the administration L-ASP catalyzed glutamine depletion with irradiated co-cultures and catalyzed tumor volume reduction in a mouse model. The clinical history of L-ASP for leukemia patients supports the viability for its repurposing as a radio-sensitizer for PCa patients

Bioelectrical regulation of cell cycle and the planarian model system.

Cell cycle regulation through the manipulation of endogenous membrane potentials offers tremendous opportunities to control cellular processes during tissue repair and cancer formation. However, the molecular mechanisms by which biophysical signals modulate the cell cycle remain underappreciated and poorly understood. Cells in complex organisms generate and maintain a constant voltage gradient across the plasma membrane known as the transmembrane potential. This potential, generated through the combined efforts of various ion transporters, pumps and channels, is known to drive a wide range of cellular processes such as cellular proliferation, migration and tissue regeneration while its deregulation can lead to tumorigenesis. These cellular regulatory events, coordinated by ionic flow, correspond to a new and exciting field termed molecular bioelectricity. We aim to present a brief discussion on the biophysical machinery involving membrane potential and the mechanisms mediating cell cycle progression and cancer transformation. Furthermore, we present the planarian Schmidtea mediterranea as a tractable model system for understanding principles behind molecular bioelectricity at both the cellular and organismal level. This article is part of a Special Issue entitled: Membrane channels and transporters in cancers

DNA damage and tissue repair: What we can learn from planaria.

Faithful renewal of aging and damaged tissues is central to organismal lifespan. Stem cells (SCs) generate the cellular progeny that replenish adult tissues across the body but this task becomes increasingly compromised over time. The age related decline in SC-mediated tissue maintenance is a multifactorial event that commonly affects genome integrity. The presence of DNA damage in SCs that are under continuous demand to divide poses a great risk for age-related disorders such as cancer. However, performing analysis of SCs with genomic instability and the DNA damage response during tissue renewal present significant challenges. Here we introduce an alternative experimental system based on the planaria flatworm Schmidtea mediterranea to address at the organismal level studies intersecting SC-mediated tissue renewal in the presence of genomic instability. Planaria have abundant SCs (neoblasts) that maintain high rates of cellular turnover and a variety of molecular tools have been developed to induce DNA damage and dissect how neoblasts respond to this stressor. S. mediterranea displays high evolutionary conservation of DNA repair mechanisms and signaling pathways regulating adult SCs. We describe genetically induced-DNA damage models and highlight body-wide signals affecting cellular decisions such as survival, proliferation, and death in the presence of genomic instability. We also discuss transcriptomic changes in the DNA damage response during injury repair and propose DNA repair as key component of tissue regeneration. Additional studies using planaria will provide insights about mechanisms regulating survival and growth of cells with DNA damage during tissue renewal and regeneration

Cancer epithelia-derived mitochondrial DNA is a targetable initiator of a paracrine signaling loop that confers taxane resistance.

Stromal-epithelial interactions dictate cancer progression and therapeutic response. Prostate cancer (PCa) cells were identified to secrete greater concentration of mitochondrial DNA (mtDNA) compared to noncancer epithelia. Based on the recognized coevolution of cancer-associated fibroblasts (CAF) with tumor progression, we tested the role of cancer-derived mtDNA in a mechanism of paracrine signaling. We found that prostatic CAF expressed DEC205, which was not expressed by normal tissue-associated fibroblasts. DEC205 is a transmembrane protein that bound mtDNA and contributed to pattern recognition by Toll-like receptor 9 (TLR9). Complement C3 was the dominant gene targeted by TLR9-induced NF-κB signaling in CAF. The subsequent maturation complement C3 maturation to anaphylatoxin C3a was dependent on PCa epithelial inhibition of catalase in CAF. In a syngeneic tissue recombination model of PCa and associated fibroblast, the antagonism of the C3a receptor and the fibroblastic knockout of TLR9 similarly resulted in immune suppression with a significant reduction in tumor progression, compared to saline-treated tumors associated with wild-type prostatic fibroblasts. Interestingly, docetaxel, a common therapy for advanced PCa, further promoted mtDNA secretion in cultured epithelia, mice, and PCa patients. The antiapoptotic signaling downstream of anaphylatoxin C3a signaling in tumor cells contributed to docetaxel resistance. The inhibition of C3a receptor sensitized PCa epithelia to docetaxel in a synergistic manner. Tumor models of human PCa epithelia with CAF expanded similarly in mice in the presence or absence of docetaxel. The combination therapy of docetaxel and C3 receptor antagonist disrupted the mtDNA/C3a paracrine loop and restored docetaxel sensitivity