Light‐Activated Transcription and Repression by Using Photocaged SERMs

Abstract

Recently developed methods to regulate the spatial and temporal patterning of genes in a light-directed manner hold promise as powerful tools for exploring the function of genes that act through their unique spatiotemporal patterning. To further explore the application of photocaged ligands of nuclear receptors to control gene expression patterning, the actions of photocaged analogues of selective estrogen-receptor modulators (SERMs) have been evaluated. Photocaged derivatives of hydroxytamoxifen (NB-Htam) and guanidine tamoxifen (NB-Gtam) have been synthesized that selectively antagonize ER alpha- and ER beta-mediated transcription at classic estrogen response elements (EREs) in response to light. When present only intracellularly, Htam and Gtam provide a similar transient repression response. When SERMs are allowed to diffuse out of the cell, transcription is recovered at a similar rate for Htam and Gtam (6.4 and 5.6 h(-1)), but is notably faster than is observed with the covalently binding SERM tamoxifen aziridine (Taz) (3.8 h(-1)). This suggests that the duration of agonist action is controlled by ligand off-rates/diffusion and not by receptor turnover. Gtam activates ER beta-mediated transcription at AP1 sites in a similar way to what has previously been reported for Htam. NB-Gtam and NB-Tam provide a light-activated transcription response at AP1-driven reporters, thus illustrating the unique ability of photocaged SERMs to simultaneously mediate light-activated transcription and repression.

Photoregulating RNA Digestion Using Azobenzene Linked Dumbbell Antisense Oligodeoxynucleotides

“…Thus, the introduction of these photoresponsive units in nucleic acid molecules has been explored and established in diverse applications, for instance, inhibition of duplex formation, control of gene transcription, regulation of aptamer recognition, manipulation of enzyme activities, and gene expression. And yet, more photoregulation strategies are focused on introduction of photolabile molecules that could be irreversibly removed by light irradiation, 8 Azobenzene has served as the excellent choice since the photoinduced reversible trans−cis isomerization of azobenzenes is accompanied by a large change in geometry and the considerable impact on structures and activities of nucleic acids. 34−36 Pioneering works related to photoswitchable nucleic acids were achieved by Komiyama, Asanuma, and co-workers.…”

Taking control of gene expression with light-activated oligonucleotides

“…These include diverse methods for focally delivering genetic material, such as by electro- or laser poration (10); the use of promoters with known temporal and tissue-specific expression profiles; diffusion of bioactive small molecules, which are typically obtained by screening large compound libraries (11); tetracycline-inducible constructs (12); Cre- lox P recombinase, which can be made inducible by heat-shock (13) or small molecules such as doxycycline, aldo-sterone, and tamoxifen (14); and light-activated (caged) compounds (3). We address the last method here, of which there are many examples: caged neurotransmitters (15,16), hormones (17,18), amino acids (1922), peptides (23,24), proteins (23,2532), calcium (33) and other second messengers such as cAMP (3437), and nitric oxide (NO) (38,39). In the caged state, these molecules are transiently blocked from performing their biological functions.…”

4-Hydroxytamoxifen probes for light-dependent spatiotemporal control of Cre-ER mediated reporter gene expression

“…[17][18][19][20][21] We previously published a study validating this concept using a photocaged tamoxifen (TAM). 22 In that study, an o-nitrobenzyl (ONB)-alkylated N-quaternary salt 15,23,24 of tamoxifen (ONB-TAM) was synthesized and found to be water soluble, and its activity was neutralized until exposed to long wave ultraviolet light (UVA, 365 nm).…”