RESUMO
Autophagy is activated in response to cellular stressors and mediates lysosomal degradation and recycling of cytoplasmic material and organelles as a temporary cell survival mechanism. Defective autophagy is implicated in human pathology, as disruption of protein and organelle homeostasis enables disease-promoting mechanisms such as toxic protein aggregation, oxidative stress, genomic damage, and inflammation. We previously showed that autophagy-defective immortalized mouse mammary epithelial cells are susceptible to metabolic stress, DNA damage, and genomic instability. We now report that autophagy deficiency is associated with endoplasmic reticulum (ER) and oxidative stress, and with deregulation of p62-mediated keratin homeostasis in mammary cells, allograft tumors, and mammary tissues from genetically engineered mice. In human breast tumors, high phospho(Ser73)-K8 levels are inversely correlated with Beclin 1 expression. Thus, autophagy preserves cellular fitness by limiting ER and oxidative stress, a function potentially important in autophagy-mediated suppression of mammary tumorigenesis. Furthermore, autophagy regulates keratin homeostasis in the mammary gland via a p62-dependent mechanism. High phospho(Ser73)-K8 expression may be a marker of autophagy functional status in breast tumors and, as such, could have therapeutic implications for breast cancer patients.
Assuntos
Autofagia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Homeostase , Queratina-8/metabolismo , Neoplasias Mamárias Experimentais/metabolismo , Neoplasias Mamárias Experimentais/patologia , Animais , Autofagia/genética , Domínio Catalítico/genética , Linhagem Celular Transformada , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Transformação Celular Neoplásica/patologia , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/patologia , Feminino , Homeostase/genética , Humanos , Queratina-8/genética , Masculino , Neoplasias Mamárias Experimentais/genética , Camundongos , Camundongos Knockout , Camundongos Nus , Camundongos Transgênicos , Estresse Oxidativo/genética , Fosfosserina/metabolismoRESUMO
Autophagy is a catabolic process involving self-digestion of cellular organelles during starvation as a means of cell survival; however, if it proceeds to completion, autophagy can lead to cell death. Autophagy is also a haploinsufficient tumor suppressor mechanism for mammary tumorigenesis, as the essential autophagy regulator beclin1 is monoallelically deleted in breast carcinomas. However, the mechanism by which autophagy suppresses breast cancer remains elusive. Here we show that allelic loss of beclin1 and defective autophagy sensitized mammary epithelial cells to metabolic stress and accelerated lumen formation in mammary acini. Autophagy defects also activated the DNA damage response in vitro and in mammary tumors in vivo, promoted gene amplification, and synergized with defective apoptosis to promote mammary tumorigenesis. Therefore, we propose that autophagy limits metabolic stress to protect the genome, and that defective autophagy increases DNA damage and genomic instability that ultimately facilitate breast cancer progression.
Assuntos
Autofagia/fisiologia , Carcinoma/patologia , Dano ao DNA/fisiologia , Neoplasias Mamárias Animais/patologia , Estresse Fisiológico/metabolismo , Animais , Proteínas Reguladoras de Apoptose , Proteína Beclina-1 , Carcinoma/genética , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Amplificação de Genes/fisiologia , Predisposição Genética para Doença , Instabilidade Genômica/fisiologia , Perda de Heterozigosidade/fisiologia , Neoplasias Mamárias Animais/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas/genética , Células Tumorais CultivadasRESUMO
Whole-cell patch clamp and polarographic oxygen partial pressure (pO2) measurements were used to establish the sensitivity of high-voltage-activated (HVA) Ca2+ channel subtypes of CA1 hippocampal neurons of rats to hypoxic conditions. Decrease of pO2 to 15-30 mm Hg induced a potentiation of HVA Ca2+ currents by 94%. Using selective blockers of N- and L-types of calcium channels, we found that inhibition of L-type channels decreased the effect by 54%, whereas N-type blocker attenuated the effect by 30%. Taking into account the ratio of currents mediated by these channel subtypes in CA1 hippocampal neurons, we concluded that both types of HVA Ca2+ channels are sensitive to hypoxia, however, L-type was about 3.5 times more sensitive to oxygen reduction.
