Laboratory of Cancer Biology and Genetics, National Cancer Institute, Bethesda, MD 20892, USA.
CCR Collaborative Bioinformatics Resource, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
Carcinogenesis. 2022 Dec 31;43(12):1149-1161. doi: 10.1093/carcin/bgac083.
Environmental and molecular carcinogenesis are linked by the discovery that chemical carcinogen induced-mutations in the Hras or Kras genes drives tumor development in mouse skin. Importantly, enhanced expression or allele amplification of the mutant Ras gene contributes to selection of initiated cells, tumor persistence, and progression. To explore the consequences of Ras oncogene signal strength, primary keratinocytes were isolated and cultured from the LSL-HrasG12D and LSL-KrasG12D C57BL/6J mouse models and the mutant allele was activated by adeno-Cre recombinase. Keratinocytes expressing one (H) or two (HH) mutant alleles of HrasG12D, one KrasG12D allele (K), or one of each (HK) were studied. All combinations of activated Ras alleles stimulated proliferation and drove transformation marker expression, but only HH and HK formed tumors. HH, HK, and K sustained long-term keratinocyte growth in vitro, while H and WT could not. RNA-Seq yielded two distinct gene expression profiles; HH, HK, and K formed one cluster while H clustered with WT. Weak MAPK activation was seen in H keratinocytes but treatment with a BRAF inhibitor enhanced MAPK signaling and facilitated tumor formation. K keratinocytes became tumorigenic when they were isolated from mice where the LSL-KrasG12D allele was backcrossed from the C57BL/6 onto the FVB/N background. All tumorigenic keratinocytes but not the non-tumorigenic precursors shared a common remodeling of matrisomal gene expression that is associated with tumor formation. Thus, RAS oncogene signal strength determines cell-autonomous changes in initiated cells that are critical for their tumor-forming potential.
环境和分子致癌学是相关的，因为发现化学致癌剂诱导的 Hras 或 Kras 基因突变驱动了小鼠皮肤的肿瘤发展。重要的是，突变 Ras 基因的增强表达或等位基因扩增有助于起始细胞的选择、肿瘤的持续存在和进展。为了探讨 Ras 癌基因信号强度的后果，从 LSL-HrasG12D 和 LSL-KrasG12D C57BL/6J 小鼠模型中分离和培养原代角质形成细胞，并通过腺病毒-Cre 重组酶激活突变等位基因。研究了表达一个（H）或两个（HH）突变等位基因的 HrasG12D、一个 KrasG12D 等位基因（K）或每个（HK）的突变等位基因的角质形成细胞。所有激活的 Ras 等位基因组合都刺激增殖并驱动转化标志物表达，但只有 HH 和 HK 形成肿瘤。HH、HK 和 K 在体外维持长期角质形成细胞生长，而 H 和 WT 则不能。RNA-Seq 产生了两个不同的基因表达谱；HH、HK 和 K 形成一个簇，而 H 与 WT 聚类。在 H 角质形成细胞中观察到弱的 MAPK 激活，但用 BRAF 抑制剂处理增强了 MAPK 信号传导并促进了肿瘤形成。当 LSL-KrasG12D 等位基因从 C57BL/6 回交至 FVB/N 背景时，从携带该等位基因的小鼠中分离出 K 角质形成细胞时，它们变得具有致瘤性。所有致瘤性角质形成细胞但不是非致瘤性前体都共享基质基因表达的共同重塑，这与肿瘤形成有关。因此，RAS 癌基因信号强度决定了起始细胞的细胞自主性变化，这对于它们的肿瘤形成潜力至关重要。
