Wolvetang Ernst J, Pera Martin F, Zuckerman Kenneth S
The Australian Stem Cell Centre and Department of Anatomy and Cell Biology, Monash University, Building 75 STRIP, Wellington Road, Clayton, Vic. 3800, Australia.
Biochem Biophys Res Commun. 2007 Nov 23;363(3):610-5. doi: 10.1016/j.bbrc.2007.09.035. Epub 2007 Sep 20.
Gap junction intracellular communication (GJIC) allows the direct transport of small molecules between adjacent cells. We hypothesized that siRNAs in one hESC could inhibit target RNA expression in another hESC via GJIC. We co-cultured green fluorescent protein (GFP)-expressing ENVY hESC with non-GFP-expressing hESC, which had been transduced to stably express shRNA directed against GFP. We discovered that the GFP shRNA expressing hESC inhibited GFP expression in the adjacent GFP-expressing hESC in a dose-dependent manner. This downregulation of GFP expression in ENVY cells was not observed when the co-cultured cells had been transduced with a non-functional GFP shRNA that was mutated at two nucleotides or when the cells were incubated with the GJIC inhibitor, alpha-glycyrrhetinic acid (alpha-GA). We conclude that 21-23 bp double-stranded shRNA/siRNA oligonucleotides are able to move through gap junctions between hESCs and thus can affect gene expression in neighbouring hESC. This novel intercellular gene expression regulatory mechanism may offer new approaches to manipulation of hESC.
间隙连接细胞内通讯（GJIC）允许小分子在相邻细胞之间直接运输。我们推测一个人胚胎干细胞（hESC）中的小干扰RNA（siRNA）可通过GJIC抑制另一个hESC中的靶RNA表达。我们将表达绿色荧光蛋白（GFP）的ENVY hESC与未表达GFP的hESC共培养，后者已被转导以稳定表达针对GFP的短发夹RNA（shRNA）。我们发现表达GFP shRNA的hESC以剂量依赖的方式抑制相邻的表达GFP的hESC中的GFP表达。当共培养的细胞用在两个核苷酸处发生突变的无功能GFP shRNA转导时，或者当细胞与GJIC抑制剂α-甘草次酸（α-GA）孵育时，未观察到ENVY细胞中GFP表达的这种下调。我们得出结论，21 - 23个碱基对的双链shRNA/siRNA寡核苷酸能够穿过hESC之间的间隙连接，因此可以影响相邻hESC中的基因表达。这种新的细胞间基因表达调控机制可能为hESC的操作提供新方法。
