Background/Objectives: The inhibitor of growth family member 3 (ING3) acts as an epigenetic reader through physical interactions with histone-modifying enzymes and subsequent chromatin remodelling processes. It is involved in various cellular functions, such as cell cycle control, cell growth, and apoptosis. Although ING3 was assigned tumour suppressor candidate status in some types of cancers, including prostate cancer, some studies suggest it acts to promote growth. To address these contradictory reports regarding its role in the initiation and progression of prostate cancer, we specifically addressed the question of whether ablation of ING3 in the mouse prostate is sufficient to initiate malignant transformation of the prostate and support its (candidate) tumour suppressor status. Methods: To generate the prostate-specificIng3knockout mouse, paternal inheritance of the PB-Cre4 transgene was used, while for the generation of a global knockout control, a female mouse harbouring the PB-Cre4 transgene was utilized. To determine the recombination efficiency of the Cre-LoxP system in the prostate at theIng3locus, a duplex probe-based digital PCR assay capable of counting undisruptedIng3copies was designed. The impact of DNA recombination on the protein level was investigated by immunohistochemical staining of prostate tissue samples. Results: In the prostate-specific knockout, digital PCR analysis revealed mosaic gene deletion. We found recombination efficiencies in the anterior, dorsolateral, and ventral prostate lobes ranging from approximately 15 to 30%. ING3 staining in the prostate was faint with no detectable differences in signal intensity between the knockout specimen and wild-type controls. This low ING3 expression in the prostate is consistent with observations of X-gal staining of anIng3-LacZ reporter allele. Immunohistochemistry showed increased expression of DNA-damage-associated markers γH2AX and 53BP1. However, no gross anatomical abnormalities or prostate intraepithelial neoplasia (PIN) lesions in the prostate of tissue-specific knockout animals compared to wild-type controls were observed. Conclusions: Altogether, our data provide evidence that disruption of ING3 expression in prostate cells does not lead to malignant transformation and challenges the idea that ING3 acts primarily in a tumour-suppressive manner. Furthermore, this work supports the crucial role of ING3 in maintaining genomic stability, and we confirmed the embryonic lethal phenotype of homozygousIng3null mice that is rescued by ectopic expression of ING3.
背景/目的：生长抑制因子家族成员3（ING3）通过与组蛋白修饰酶的物理相互作用及后续的染色质重塑过程，发挥表观遗传阅读器功能。它参与多种细胞功能，如细胞周期调控、细胞生长和凋亡。尽管ING3在前列腺癌等某些癌症中被列为候选抑癌基因，但部分研究提示其具有促生长作用。为澄清其在前列腺癌发生发展中的矛盾报道，本研究重点探讨小鼠前列腺中ING3的缺失是否足以启动前列腺恶性转化，从而验证其（候选）抑癌基因地位。方法：通过父系遗传PB-Cre4转基因构建前列腺特异性Ing3敲除小鼠，同时利用携带PB-Cre4转基因的雌鼠建立全身性敲除对照。为检测Cre-LoxP系统在前列腺Ing3位点的重组效率，设计了基于双探针的数字PCR检测方法以计数未断裂的Ing3拷贝。通过前列腺组织样本的免疫组化染色研究DNA重组对蛋白水平的影响。结果：数字PCR分析显示前列腺特异性敲除小鼠存在嵌合性基因缺失，其前列腺前叶、背外侧叶和腹叶的重组效率约为15%至30%。前列腺组织中ING3染色微弱，敲除样本与野生型对照的信号强度无显著差异。这种低表达现象与Ing3-LacZ报告等位基因的X-gal染色观察结果一致。免疫组化检测显示DNA损伤标志物γH2AX和53BP1表达增加，但与野生型对照相比，组织特异性敲除动物未发现明显解剖结构异常或前列腺上皮内瘤变（PIN）病变。结论：本研究数据表明，前列腺细胞中ING3表达的破坏不会导致恶性转化，这对ING3主要发挥抑癌作用的观点提出了挑战。此外，本研究证实了ING3在维持基因组稳定性中的关键作用，并通过ING3异位表达成功挽救了纯合Ing3缺失小鼠的胚胎致死表型。
Loss of ING3 in the Prostate Leads to Activation of DNA Damage Repair Markers
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