Differential Reprogramming of Isogenic Colorectal Cancer Cells by Distinct Activating KRAS Mutations

Oncogenic
mutations of Ras at codons 12, 13, or 61, that render
the protein constitutively active, are found in ∼16% of all
cancer cases. Among the three major Ras isoforms, KRAS is the most
frequently mutated isoform in cancer. Each Ras isoform and tumor type
displays a distinct pattern of codon-specific mutations. In colon
cancer, KRAS is typically mutated at codon 12, but a significant fraction
of patients have mutations at codon 13. Clinical data suggest different
outcomes and responsiveness to treatment between these two groups.
To investigate the differential effects upon cell status associated
with KRAS mutations we performed a quantitative analysis of the proteome
and phosphoproteome of isogenic SW48 colon cancer cell lines in which
one allele of the endogenous gene has been edited to harbor specific
KRAS mutations (G12V, G12D, or G13D). Each mutation generates a distinct
signature, with the most variability seen between G13D and the codon
12 KRAS mutants. One notable example of specific up-regulation in
KRAS codon 12 mutant SW48 cells is provided by the short form of the
colon cancer stem cell marker doublecortin-like Kinase 1 (DCLK1) that
can be reversed by suppression of KRAS.

致癌突变（oncogenic mutation）在Ras蛋白（Ras）的密码子12、13或61位发生时，可使该蛋白持续激活，这类突变在所有癌症病例中占比约16%。在Ras家族的三种主要同工型（isoform）中，KRAS是癌症中突变频率最高的同工型。每种Ras同工型与对应的肿瘤类型均呈现出独特的密码子特异性突变模式。在结肠癌中，KRAS通常在密码子12位发生突变，但仍有相当比例的患者其KRAS突变位点位于密码子13位。临床数据显示，这两类患者的预后及治疗响应性存在显著差异。为探究KRAS突变对细胞状态的差异化影响，我们对同基因SW48结肠癌细胞系的蛋白质组（proteome）与磷酸化蛋白质组（phosphoproteome）进行了定量分析。该细胞系的内源KRAS基因单等位基因（allele）已被编辑为携带特定突变（G12V、G12D或G13D）的形式。每种KRAS突变均可产生独特的分子特征谱，其中G13D突变体与密码子12位KRAS突变体之间的差异最为显著。值得注意的是，在密码子12位突变的SW48细胞中，结肠癌干细胞标志物双皮质素样激酶1（DCLK1）的短亚型表达显著上调，且该上调效应可通过抑制KRAS逆转。