Arid1a restrains Kras-dependent changes in acinar cell identity

Mutations in members of the SWI/SNF chromatin remodeling family are common events in human cancer, but the mechanisms whereby disruption of SWI/SNF components alters tumorigenesis remain poorly understood. To model the effect of loss of function mutations in the SWI/SNF subunit Arid1a in pancreatic ductal adenocarcinoma (PDAC) initiation, we directed shRNA triggered, inducible and reversible suppression of Arid1a to the pancreas of mice in the setting of oncogenic KrasG12D. Arid1a cooperates with Kras in the adult pancreas as postnatal silencing of Arid1a following sustained KrasG12D expression induces rapid and irreversible reprogramming of acinar cells into mucinous PDAC precursor lesions. In contrast, Arid1a silencing during embryogenesis, concurrent with KrasG12D activation, leads to retention of acinar cell fate. Together, our results demonstrate Arid1a as a critical modulator of Kras-dependent changes in acinar cell identity, and underscore an unanticipated influence of timing and genetic context on the effects of SWI/SNF complex alterations in epithelial tumorigenesis. Overall design: Identification of the transcritpional changes induced upon dox-inducible Arid1a knockdown in mutant Kras pancreatic epithelial cells: Arid1a-downregulated vs Arid1a-expressing mutant Kras-pancreatic epithelial cells were FACS-sorted from pancreata from p48Cre;RIK;LSL-KrasG12;TRE-shRNA mice (harbouring Arid1a shRNAs -6421 or 1803- or control Renilla shRNA -713-) at day 5 post- doxycycline treatment. Mice were placed on doxycycline diet at 5 weeks of age to induce expression of the indicated shRNAs. 3 biological replicates (independent mice) were used per shRNA condition (713, 6421, 1803).SWI/SNF染色质重塑家族（SWI/SNF chromatin remodeling family）成员的突变在人类癌症中十分常见，但SWI/SNF组分失调改变肿瘤发生的具体机制仍不甚明晰。为构建SWI/SNF亚基Arid1a功能缺失突变在胰腺导管腺癌（PDAC，pancreatic ductal adenocarcinoma）起始阶段的疾病模型，我们在致癌性KrasG12D背景下，向小鼠胰腺中导入短发夹RNA（shRNA）介导的可诱导且可逆的Arid1a抑制体系。 Arid1a与成年小鼠胰腺中的Kras存在协同效应：在持续表达KrasG12D后对Arid1a进行出生后沉默，可快速且不可逆地将腺泡细胞重编程为黏液性胰腺癌前病变。与之相反，在胚胎发生期伴随KrasG12D激活时沉默Arid1a，则会保留腺泡细胞的细胞身份。综上，本研究结果证实Arid1a是Kras依赖的腺泡细胞身份改变的关键调控因子，并凸显了时序与遗传背景对SWI/SNF复合物失调在上皮肿瘤发生中作用的未曾预料到的影响。 整体实验设计：鉴定多西环素（doxycycline，dox）诱导的Arid1a敲低对携带突变Kras的胰腺上皮细胞所引发的转录变化：将经多西环素处理5天后的p48Cre;RIK;LSL-KrasG12;TRE-shRNA小鼠（分别携带Arid1a shRNA-6421、Arid1a shRNA-1803或对照海肾shRNA-713）的胰腺组织进行荧光激活细胞分选（FACS），分离得到Arid1a下调组与Arid1a表达组的突变Kras胰腺上皮细胞。实验小鼠于5周龄时饲喂多西环素饲料以诱导指定shRNA的表达。每个shRNA处理组（713、6421、1803）均设置3例生物学重复（独立小鼠样本）。