There is a desire to engineer mammalian host cell lines to improve cell growth/biomass accumulation and recombinant biopharmaceutical protein production in industrially relevant cell lines such as the CHOK1 and HEK293 cell lines. The over-expression of individual subunits of the eukaryotic translation factor eIF3 in mammalian cells has previously been shown to result in oncogenic properties being imparted on cells, including increased cell proliferation and growth and enhanced global protein synthesis rates. Here we report on the engineering of CHOK1 and HEK cells to over-express the eIF3i and eIF3c subunits of the eIF3 complex and the resultant impact on cell growth and a reporter of exogenous recombinant protein production. Transient over-expression of eIF3i in HEK293 and CHOK1 cells resulted in a modest increase in total eIF3i amounts (maximum 40% increase above control) and an approximate 10% increase in global protein synthesis rates in CHOK1 cells. Stable over-expression of eIF3i in CHOK1 cells was not achievable, most likely due to the already high levels of eIF3i in CHO cells compared to HEK293 cells, but was achieved in HEK293 cells. HEK293 cells engineered to over-express eIF3i had faster growth that was associated with increased c-Myc expression, achieved higher cell biomass and gave enhanced yields of a reporter of recombinant protein production. Whilst CHOK1 cells could not be engineered to over-express eIF3i directly, they could be engineered to over-express eIF3c, which resulted in a subsequent increase in eIF3i amounts and c-Myc expression. The CHOK1 eIF3c engineered cells grew to higher cell numbers and had enhanced cap- and IRES-dependent recombinant protein synthesis. Collectively these data show that engineering of subunits of the eIF3 complex can enhance cell growth and recombinant protein synthesis in mammalian cells in a cell specific manner that has implications for the engineering or selection of fast growing or high producing cells for production of recombinant proteins.

需要对哺乳动物宿主细胞系进行工程改造，以改善工业相关细胞系如 CHOK1 和 HEK293 细胞系中的细胞生长/生物量积累和重组生物药物蛋白的生产。哺乳动物细胞中真核翻译因子 eIF3 的单个亚基的过度表达先前已被证明会导致细胞具有致癌特性，包括增加细胞增殖和生长以及提高整体蛋白质合成率。在这里，我们报告了 CHOK1 和 HEK 细胞的工程，以过度表达 eIF3 复合物的 eIF3i 和 eIF3c 亚基，以及由此产生的对细胞生长的影响和外源重组蛋白生产的报告。HEK293 和 CHOK1 细胞中 eIF3i 的瞬时过表达导致总 eIF3i 量适度增加（比对照最多增加 40%），并且 CHOK1 细胞中的整体蛋白质合成率增加约 10%。与 HEK293 细胞相比，CHOK1 细胞中 eIF3i 的稳定过表达无法实现，这很可能是由于 CHO 细胞中 eIF3i 的水平已经很高，但在 HEK293 细胞中实现了。为过度表达 eIF3i 而设计的 HEK293 细胞具有更快的生长，这与 c-Myc 表达增加有关，实现了更高的细胞生物量并提高了重组蛋白生产报告基因的产量。虽然 CHOK1 细胞不能被设计为直接过度表达 eIF3i，但它们可以被设计为过度表达 eIF3c，这导致随后 eIF3i 数量和 c-Myc 表达的增加。CHOK1 eIF3c 工程细胞生长到更高的细胞数量，并增强了帽和 IRES 依赖性重组蛋白合成。总的来说，这些数据表明，工程化 eIF3 复合物的亚基可以以细胞特异性方式增强哺乳动物细胞中的细胞生长和重组蛋白质合成，这对工程化或选择快速生长或高产细胞以生产重组蛋白质具有影响。