Terskikh VV et al. (2012) Stem Cell Rev "Label retaining cells and cutaneous stem cells."

Vorotelyak EA, Terskikh VV, Vasiliev AV

[Stem Cell Rev, 2012]

This is a comprehensive review on label retaining cells (LRC) in epidermal development and homeostasis. The precise in vivo identification and location of epidermal stem cells is a crucial issue in cutaneous biology. We discuss here the following problems: (1) Identification and location of LRC in the interfollicular epithelium and hair follicle; (2) The proliferative potential of LRC and their role in cutaneous homeostasis (3); LRC phenomenon and the Immortal Strand Hypothesis, which suggests an alternative mechanism for retention of genetic information; (4) Significance of LRC studies for development of stem cell concept. Now, it seems evident that LRC are a frequent feature of stem cell niches and revealing highly dormant LRC may be used for identification of stem cell niches in different tissues. LRC were used for screening specific markers of epidermal stem cells. Within a given tissue stem cells have different proliferative characteristics. There are more frequently cycling stem cells which function primarily in homeostasis, while LRC form a reserve of dormant, may be ultimate, stem cells, which are set aside for regeneration of injury or unforeseen need. The authors suggest that LRC dormancy described in Mammalia has much in common with developmental quiescence found in some other animals. For example in C. elegans reproductive system, vulval precursor cells have developmentally programmed cell-cycle arrest in the first larval stage, and then undergo an extended period of quiescence before resuming proliferation. Another example of developmental quiescence is the diapause, a widespread phenomenon exhibited by animals ranging from nematodes to mammals, often occurring at genetically predetermined life history stage.

Terskikh A et al. (2000) Science ""Fluorescent timer": protein that changes color with time."

Matz M, Ermakova G, Siebert P, Kim SK, Lukyanov S, Kajava AV, Weissman I, Zaraisky A, Terskikh A, Tan PBO, Fradkov A, Zhao X

[Science, 2000]

We generated a mutant of the red fluorescent protein drFP583. The mutant (E5) changes its fluorescence from green to red over time. The rate of color conversion is independent of protein concentration and therefore can be used to trace time-dependent expression. We used in vivo labeling with E5 to measure expression from the heat shock-dependent promoter in Caenorhabditis elegans and from the Otx-2 promoter in developing Xenopus embryos. Thus, E5 is a "fluorescent timer" that can be used to monitor both activation and down-regulation of target promoters on the whole-organism scale.AD - School of Medicine, Stanford University, Stanford, CA 94305, USA. Alexey.Terskikh@Stanford.eduFAU - Terskikh, AAU - Terskikh AFAU - Fradkov, AAU - Fradkov AFAU - Ermakova, GAU - Ermakova GFAU - Zaraisky, AAU - Zaraisky AFAU - Tan, PAU - Tan PFAU - Kajava, A VAU - Kajava AVFAU - Zhao, XAU - Zhao XFAU - Lukyanov, SAU - Lukyanov SFAU - Matz, MAU - Matz MFAU - Kim, SAU - Kim SFAU - Weissman, IAU - Weissman IFAU - Siebert, PAU - Siebert PLA - engID - 1 RO3 TW01362-01/TW/FICPT - Journal ArticleCY - UNITED STATESTA - ScienceJID - 0404511RN - 0 (Heat-Shock Proteins)RN - 0 (Luminescent Proteins)RN - 0 (Nerve Tissue Proteins)RN - 0 (Otx2 protein)RN - 0 (Trans-Activators)RN - 0 (red fluorescent protein)SB - IM

Allan Mah et al. (2005) International Worm Meeting "Assessment of Fluorescent Timer as a tool for assaying transcriptional regulation in C. elegans"

Derek Chew, Robert Johnsen, David Baillie, Domena Tu, Allan Mah

[International Worm Meeting, 2005]

Fluorescent Timer, a proprietary fluorescent reporter from Clontech, has the unique property of shifting its emission fluorescence from green (480nm) to red (583nm) in a time/intramolecular reaction dependent manner. Red fluorescence was documented to occur two to four hours after initial green fluorescence is detected (Terskikh et al. 2000). I have attempted to use this reporter to assay for message turnover under the control of various upstream regulatory regions. Regulatory regions were selected on the basis of showing limited temporal expression during early development and/or in various tissues to test the efficacy of the reporter as a general assay of transcriptional regulation. Regulatory regions were linked to the reporter via PCR fusion (Hobert, O. 2002). All transgenic lines containing upstream regulatory regions fused to the raw reporter failed to produce fluorescence. Upon the addition of a 3UTR and the insertion of two adenosine residues directly upstream of the translational start site, 10/14 transgenic lines displayed fluorescence. Embryonic expression, which was observed using the same upstream regulatory regions fused to GFP, was not observed in any of the Fluorescent Timer reporter lines constructed. To possibly counter to effects of cytoplasmic toxicity of the Fluorescent Timer protein, and to concentrate the signal to the nucleus, I fused a 5NLS to the construct. Unexpectedly, only 3/14 lines displayed fluorescence leading to the assumption that Fluorescent Timer may be more toxic when concentrated to the nucleus. This result seems to agree with the results that no Fluorescent Timer expression was observed during early development possibly due to toxicity of the construct. Fluorescent Timer may be a useful system for the assay of temporal regulation of transcription during later stages of C. elegans development. References: Terskikh A, Fradkov A, Ermakova G, Zaraisky A, Tan P, Kajava AV, Zhao X, Lukyanov S, Matz M, Kim S, Weissman I, Siebert P. 2000, "Fluorescent timer": protein that changes color with time. Science. Nov 24;290(5496):1585-8. Hobert O. 2002, PCR fusion-based approach to create reporter gene constructs for expression analysis in transgenic C. elegans. Biotechniques. Apr;32(4):728-30.