A transcriptional variant of the LC3A gene is involved in autophagy and frequently inactivated in human cancers

Abstract

Microtubule-associated protein 1 light chain 3 has an important role in autophagy. The human LC3 gene family has five members, LC3A (variant-1: v1 and -2: v2), LC3B, LC3B2 and LC3C. Although a form of LC3B modified by phosphatidylethanolamine (form-II) is localized in autophagosomes, it is not clear whether other LC3 proteins also function in autophagy. Here, we examined the association between autophagy and human LC3 proteins during starvation-or p53-induced autophagy in Saos-2 cells. In an analysis of the intracellular distribution of each LC3 protein fused with GFP, GFP-LC3Av1 was frequently localized in autophagosomes with a punctate pattern, similar to GFP-LC3B. Further, endogenous LC3Av1 generated form-II and mostly localized in LC3B-positive autophagosomes during the induced autophagy. Interestingly, LC3Av1, not LC3B, was frequently inactivated at the transcriptional level in various human cancer cell lines (111/244 cell lines, 45.5%) and its inactivation was due to aberrant DNA methylation in esophageal squamous cell carcinoma (ESCC) cell lines and primary tumors. Restoration of LC3Av1 expression in KYSE170 cells, an LC3Av1-inactivated ESCC cell line, showed the inhibition of tumor growth in vivo. These results suggest that LC3Av1, not only LC3B, functions in autophagy and further, LC3Av1 may be crucial in carcinogenesis.

Raf/MEK/ERK can regulate cellular levels of LC3B and SQSTM1/p62 at expression levels

“…While LC3B has been known as the major LC3 in the autophagosome, a recent study demonstrated that the other LC3 family member LC3A-vareint1 can also mediate autophagosome formation as efficiently as LC3B [26]. Contrary to LC3B regulation, Raf activation significantly decreased LC3A-vareint1 mRNA levels (Fig.…”

“…We determined the effect of Raf/MEK/ERK activation on LC3B, a major LC3 in the autophagosome [26], and SQSTM1 in different cell types, including the human prostate cancer line LNCaP, immortalized human kidney epithelial cell line HEK293, primary cultured normal foreskin fibroblast BJ, and immortalized lung fibroblast IMR90E1A, which have been used as models to study the mechanisms underlying cellular responses to aberrant Raf/MEK/ERK activation [6,18,27]. To specifically control the pathway activity, we used the tamoxifen-regulated ΔRaf-1:ER, a CR3 catalytic domain of Raf-1 fused to hormone binding domain of the estrogen receptor [28].…”

Enhanced autophagy is required for survival in EGFR-independent EGFR-mutant lung adenocarcinoma cells

“…The core molecular machinery required for the formation of an autophagosome comprises a heterogeneous group of proteins encoded by autophagy-related ATG genes, such as ATG5, Beclin1 (also known as ATG6), and microtubule-associated protein 1 light chain 3 (LC3), a human homolog of yeast ATG8. [10][11][12] The conversion of cytosolic LC3-I to autophagosome-bound LC3-II acts as a biochemical marker of active autophagy, [10][11][12][13][14][15][16] and it has been increasingly appreciated that the immunohistochemical detection of cytoplasmic LC3 puncta in formalin-fixed, paraffinembedded specimens (FFPE) correlates with the amount of autophagosomes or autophagolysosomes. 14,17,18 Although it is known that cancer cells with Ras mutations are addicted to upregulated autophagic flux, 10,11,13,14 little has been done to clarify the role of autophagy in EGFR-driven carcinoma cells.…”

Epigenetic modifications as regulatory elements of autophagy in cancer

“…Recently, Bai et al found that the function of GFP-LC3Av1 (a LC3A variant) was similar to GFP-LC3B during the process of autophagy. In the further study, LC3Av1 was shown to be frequently methylated in ESCC and the restoration of LC3Av1 expression exerted its anti-proliferative activity in vivo [20]. The expression of autophagy-related protein Beclin 1 was significantly downregulated in breast tumors due to aberrant promoter methylation [21].…”