Isoform-specific induction of a retinoid-responsive antigen after biolistic transfection of chimaeric retinoic acid/thyroid hormone receptors into a regenerating limb.
Retinoic acid (RA) induces secretory differentiation in the wound epidermis of a regenerating amphibian limb. We investigated the role of individual RA receptor (RAR) types in the newt wound epidermis by introducing chimaeric RA/thyroid hormone (T3) receptors (chi alpha 1 and chi delta 1) that can be activated by T3. A biolistic particle delivery system was employed to transfect cells in the wound epidermis of a regenerating limb and approximately 10% of the cells in targeted surface areas expressed marker genes. Both chi alpha 1 and chi delta 1 were comparable in their ability to stimulate transcription of a synthetic reporter construct through a RA response element after activation with T3 in situ. This activation was also comparable to that obtained by the endogenous complement of RARs in the RA-treated, transfected wound epidermis. The RA-inducible WE3 antigen, a marker for secretory differentiation, which distinguishes the wound epidermis from normal skin (Tassava, R. A., Johnson-Wint, B. and Gross, J. 1986, J. Exp. Zool. 239, 229-240), was used to assess the functional role of chi alpha 1 and chi delta 1. Chimaeric receptors were transfected with an alkaline phosphatase marker gene, activated with T3, and the expression of both the marker and WE3 was analyzed by double-label immunofluorescence. Newt limbs transfected with chi delta 1 showed many double-labelled cells dependent on the presence of T3, whereas contralateral limbs transfected with an alkaline phosphatase marker lacking chimaeric receptor sequences did not.(ABSTRACT TRUNCATED AT 250 WORDS)
Pecorino
L T
LT
Ludwig Institute for Cancer Research, University College London, UK.
Lo
D C
DC
Brockes
J P
JP
D000818:Animals; D015415:Biomarkers / Q000032:analysis; D004817:Epidermis / Q000502:physiology*; D004848:Epithelium / Q000502:physiology; D005121:Extremities / Q000502:physiology*; D006023:Glycoproteins / Q000032:analysis; D007150:Immunohistochemistry; D036341:Intercellular Signaling Peptides and Proteins; D007536:Isomerism; D018168:Receptors, Retinoic Acid / Q000502:physiology*; D011988:Receptors, Thyroid Hormone / Q000502:physiology; D012038:Regeneration / Q000502:physiology*; D012455:Salamandridae / Q000502:physiology*; D014162:Transfection
D015415:Biomarkers; D006023:Glycoproteins; D036341:Intercellular Signaling Peptides and Proteins; D018168:Receptors, Retinoic Acid; D011988:Receptors, Thyroid Hormone; C086638:WE3 protein, newt