Dynamics of stimulus-expression coupling as revealed by monitoring of prolactin promoter-driven reporter activity in individual, living mammotropes.
Single-cell paradigms have greatly expanded our knowledge about stimulus-secretion coupling, but the understanding of stimulus-gene expression coupling has lagged behind for lack of a dynamic model sufficiently sensitive to provide single-cell resolution. In the present study, we made continuous indirect measurements within individual, living cells of expression dynamics both before and after treatment with a gene-activating secretagogue. This was accomplished by transfecting (via microinjection) individual, primary mammotropes with a PRL promoter-driven luciferase reporter plasmid, and then quantifying the rate of photonic emissions (reflective of endogenous gene activity). We found that individual cells exhibit spontaneous, random, short-term fluctuations of basal reporter activity and are extremely heterogeneous in terms of responses to a stimulatory agent (TRH). In addition, we found that responses are affected by several factors including the secretory status of the pituitary donor, the manner in which the stimulus is presented, and by the initial level of reporter activity. Moreover, the responsiveness of an individual cell can fluctuate dramatically over time. These results invite speculation that a given cell can "sense" its gene activation state and regulate its response accordingly to satisfy requirements for the corresponding secretory product.
Villalobos
C
C
Department of Cell Biology and Anatomy, Medical University of South Carolina, Charleston 29425, USA.
Faught
W J
WJ
Frawley
L S
LS
D000818:Animals; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D007774:Lactation; D008156:Luciferases / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D008297:Male; D008322:Mammals; D017785:Photons; D010902:Pituitary Gland / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D011388:Prolactin / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D051381:Rats; D017207:Rats, Sprague-Dawley; D012737:Sex Factors; D013973:Thyrotropin-Releasing Hormone / Q000378:metabolism / Q000494:pharmacology*; D013997:Time Factors; D015533:Transcriptional Activation; D014162:Transfection
D013973:Thyrotropin-Releasing Hormone; D011388:Prolactin; D008156:Luciferases