Induction of pluripotency by protein gain of function in cloned mouse embryos
Cell Biology
Final Report Abstract
Using SCNT, confocal microscopy, transcriptome analysis by microarray, and protein analysis by LC-MS/MS as core technologies, the Boiani group studied the natural reprogramming ability of the MII mouse oocyte. This was relevant not only to the topic of SPP 1356, but also to developmental totipotency and to oocyte biology in general. Results obtained in the SPP 1356 can be summarized in four points: 1) The natural factors that are in charge of oocyte-mediated reprogrammming, called collectively the ‘reprogrammome,’ are, in part, distinct from the reprogramming factors known from other reprogramming systems e.g. Oct4; 2) the reprogramming factors of one ooplasm are sufficient for more than one somatic nucleus; 3) the cell cycle and, in particular, DNA replication is an essential co-factor in oocyte-mediated reprogramming; and 4) reprogramming unfolds into at least the third cell cycle after SCNT and requires culture conditions as a permissive parameter. To our surprise, the key transcription factor Oct4 only allowed for marginal improvement of reprogramming rates after artificial increase of Oct4 mRNA in oocytes prior to SCNT. Given these results, where to go from here? Ongoing research by the Boiani group aims to investigate the composition of the mouse ooplasm deeper in order to pinpoint the reprogramming factors, even though these factors may not be distinct from those that process the sperm nucleus and that direct development after fertilization. For this reason, the study of cloned embryos is meaningful also for the understanding of normal development. A new project called “A forward genetics approach to the ‘reprogrammome’ is in progress to identify the protein factors whose abundance may correlate with the variable reprogramming phenotypes of oocytes from different inbred strains of mice (C3H/HeN, C57Bl/6J, DBA/2J, and 129/Sv). The oocytes and embryos of inbred strains offer the advantage of a defined genetic composition, which undoubtedly will simplify the interpretation of experimental results.
Publications
- (2010). Enhancing somatic nuclear reprogramming by Oct4 gain-of-function in cloned mouse embryos. Int J Dev Biol. 54:1649-1657
Pfeiffer MJ, Balbach ST, Esteves TC, Crosetto N, Boiani M
(See online at https://doi.org/10.1387/ijdb.103197mp) - (2010). Somatic cell nuclear reprogramming of mouse oocytes endures beyond reproductive decline. Aging Cell 10:80-95
Esteves TC, Balbach ST, Pfeiffer MJ, Araúzo-Bravo MJ, Klein DC, Sinn M, Boiani M
(See online at https://doi.org/10.1111/j.1474-9726.2010.00644.x) - (2011). 5-Hydroxymethylcytosine in the mammalian zygote is linked with epigenetic reprogramming. Nature Communication 2:241
Wossidlo M, Nakamura T, Lepikhov K, Marques CJ, Zakhartchenko V, Boiani M, Arand J, Nakano T, Reik W, Walter J
(See online at https://doi.org/10.1038/ncomms1240) - (2011). Proteomic analysis of mouse oocytes reveals 28 candidate factors of the „reprogrammome". J Proteome Research 10:2140-2153
Pfeiffer MJ, Siatkowski M, Paudel Y, Balbach ST, Baeumer N, Crosetto N, Drexler HC, Fuellen G, Boiani M
(See online at https://doi.org/10.1021/pr100706k) - (2013). Reprogramming of two somatic nuclei in the same ooplasm leads to pluripotent embryonic stem cells. Stem Cells 31(11):2343-53
Pfeiffer MJ, Esteves TC, Balbach ST, Araúzo-Bravo MJ, Stehling M, Jauch A, Houghton FD, Schwarzer C, Boiani M
(See online at https://doi.org/10.1002/stem.1497) - (2014). DNA replication is an essential component of the mouse oocyte’s reprogramming machinery. PLoS One 9(5):e97199
Wang B, Pfeiffer MJ, Schwarzer C, Arauzo-Bravo MJ, Boiani M
(See online at https://doi.org/10.1371/journal.pone.0097199) - (2014). Maternal age effect on mouse oocytes: new biological insight from proteomic analysis. Reproduction 148(1):55-72
Schwarzer C, Siatkowski M, Pfeiffer MJ, Baeumer N, Drexler HC, Wang B, Fuellen G, Boiani M
(See online at https://doi.org/10.1530/REP-14-0126) - Differences in embryo quality are associated with differences in oocyte composition: A proteomic study in inbred mice. Proteomics,
2014
Nov
4
Pfeiffer MJ, Taher L, Drexler H, Suzuki Y, Makałowski W, Schwarzer C, Wang B, Fuellen G, Boiani M
(See online at https://doi.org/10.1002/pmic.201400334)
