Reprogramming triggers endogenous L1 and Alu retrotransposition in human induced pluripotent stem cells.

Authors:
Klawitter S, Fuchs NV, Upton KR, Muñoz-Lopez M, Shukla R, Wang J, Garcia-Cañadas M, Lopez-Ruiz C, Gerhardt DJ, Sebe A, Grabundzija I, Merkert S, Gerdes P Pulgarin JA, Bock A, Held U, Witthuhn A, Haase A, Sarkadi B, Löwer J, Wolvetang EJ, Martin U, Ivics Z, Izsvák Z, Garcia-Perez JL, Faulkner GJ, Schumann GG
In:
Source: Nat Commun.
Publication Date: (2016)
Issue: 7: 10286
Research Area:
Stem Cells
Basic Research
Cells used in publication:
HFF-1
Species: human
Tissue Origin: dermal
Platform:
Nucleofector® I/II/2b
Experiment

 HFF-1 were reprogrammed by using Sleeping Beauty transposon-based plasmids pT2-OSKM or pT2-OSKML.  4x10^5 cells per well were nucleofeted with 2 mg of transposon plasmid and 0.2 mg of CMV- SB100X vector.

Abstract

Human induced pluripotent stem cells (hiPSCs) are capable of unlimited proliferation and can differentiate in vitro to generate derivatives of the three primary germ layers. Genetic and epigenetic abnormalities have been reported by Wissing and colleagues to occur during hiPSC derivation, including mobilization of engineered LINE-1 (L1) retrotransposons. However, incidence and functional impact of endogenous retrotransposition in hiPSCs are yet to be established. Here we apply retrotransposon capture sequencing to eight hiPSC lines and three human embryonic stem cell (hESC) lines, revealing endogenous L1, Alu and SINE-VNTR-Alu (SVA) mobilization during reprogramming and pluripotent stem cell cultivation. Surprisingly, 4/7 de novo L1 insertions are full length and 6/11 retrotransposition events occurred in protein-coding genes expressed in pluripotent stem cells. We further demonstrate that an intronic L1 insertion in the CADPS2 gene is acquired during hiPSC cultivation and disrupts CADPS2 expression. These experiments elucidate endogenous retrotransposition, and its potential consequences, in hiPSCs and hESCs.