TY - JOUR
T1 - Highly efficient reprogrammable mouse lines with integrated reporters to track the route to pluripotency
AU - Elbaz, Judith
AU - Puri, Mira C.
AU - Faiz, Maryam
AU - Bang, K. W.Annie
AU - Nguyen, Lena
AU - Makovoz, Bar
AU - Gertsenstein, Marina
AU - Hussein, Samer M.I.
AU - Zandstra, Peter W.
AU - Briollais, Laurent
AU - Shakiba, Nika
AU - Nagy, Andras
N1 - Funding Information:
ACKNOWLEDGMENTS. We acknowledge the contribution of the Model Production Core at The Centre for Phenogenomics for technical support in generating chimeras. We are grateful to Malgosia Kownacka for the preparation of the mouse embryonic feeders, to Balazs Varga for his help with immunofluorescence, tissue culture, and scientific discussions, to Maria Shutova for support with bioinformatics, to Qin Liang and Kristina Nagy (K.N.) for their contribution to the site of insertion detection, to Claudio Monetti (C.M.) and Julia Nakanishi for artwork editing, K.N. and C.M. for comments on the manuscript, to Rebecca Cowling for help with mouse genotyping and microscopy, to Christina Dalrymple for her help with the mouse colony,and to Dr.Yitzhak Reizel for fruitful discussions regarding the epigenetic analysis in our study. This work was supported by an Ontario Research Fund Genome and Life Sciences (Round 2) Program grant from the Ontario Ministry of Research and Innovation (to A.N., GL2-01-028), the Canada Research Chairs Program (to A.N.), and a CIHR Foundation Program Grant (to A.N.).
Publisher Copyright:
Copyright © 2022 the Author(s).
PY - 2022/12/6
Y1 - 2022/12/6
N2 - Revealing the molecular events associated with reprogramming different somatic cell types to pluripotency is critical for understanding the characteristics of induced pluripotent stem cell (iPSC) therapeutic derivatives. Inducible reprogramming factor transgenic cells or animals—designated as secondary (2°) reprogramming systems—not only provide excellent experimental tools for such studies but also offer a strategy to study the variances in cellular reprogramming outcomes due to different in vitro and in vivo environments. To make such studies less cumbersome, it is desirable to have a variety of efficient reprogrammable mouse systems to induce successful mass reprogramming in somatic cell types. Here, we report the development of two transgenic mouse lines from which 2° cells reprogram with unprecedented efficiency. These systems were derived by exposing primary reprogramming cells containing doxycycline-inducible Yamanaka factor expression to a transient interruption in transgene expression, resulting in selection for a subset of clones with robust transgene response. These systems also include reporter genes enabling easy readout of endogenous Oct4 activation (GFP), indicative of pluripotency, and reprogramming transgene expression (mCherry). Notably, somatic cells derived from various fetal and adult tissues from these 2° mouse lines gave rise to highly efficient and rapid reprogramming, with transgene-independent iPSC colonies emerging as early as 1 wk after induction. These mouse lines serve as a powerful tool to explore sources of variability in reprogramming and the mechanistic underpinnings of efficient reprogramming systems.
AB - Revealing the molecular events associated with reprogramming different somatic cell types to pluripotency is critical for understanding the characteristics of induced pluripotent stem cell (iPSC) therapeutic derivatives. Inducible reprogramming factor transgenic cells or animals—designated as secondary (2°) reprogramming systems—not only provide excellent experimental tools for such studies but also offer a strategy to study the variances in cellular reprogramming outcomes due to different in vitro and in vivo environments. To make such studies less cumbersome, it is desirable to have a variety of efficient reprogrammable mouse systems to induce successful mass reprogramming in somatic cell types. Here, we report the development of two transgenic mouse lines from which 2° cells reprogram with unprecedented efficiency. These systems were derived by exposing primary reprogramming cells containing doxycycline-inducible Yamanaka factor expression to a transient interruption in transgene expression, resulting in selection for a subset of clones with robust transgene response. These systems also include reporter genes enabling easy readout of endogenous Oct4 activation (GFP), indicative of pluripotency, and reprogramming transgene expression (mCherry). Notably, somatic cells derived from various fetal and adult tissues from these 2° mouse lines gave rise to highly efficient and rapid reprogramming, with transgene-independent iPSC colonies emerging as early as 1 wk after induction. These mouse lines serve as a powerful tool to explore sources of variability in reprogramming and the mechanistic underpinnings of efficient reprogramming systems.
KW - induced pluripotent stem cells
KW - reprogrammable mouse
KW - secondary reprogramming
KW - somatic cell reprogramming
KW - transgenic mouse
UR - http://www.scopus.com/inward/record.url?scp=85143610058&partnerID=8YFLogxK
U2 - 10.1073/pnas.2207824119
DO - 10.1073/pnas.2207824119
M3 - Article
AN - SCOPUS:85143610058
SN - 0027-8424
VL - 119
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 49
M1 - e2207824119
ER -