What is the role of the promoter in cell ablation studies?

[Eagle9]

In Chemistry and Biology the term “Promoter” has got two meanings, first:

1. (Chemistry) a substance added in small amounts to a catalyst to increase its activity

2. (Genetics) a sequence of nucleotides, associated with a structural gene, that must bind with messenger RNA polymerase before transcription can proceed

In this paper Glial Cells and Their Function in the Adult Brain: A Journey through the History of Their Ablation

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5303749/

the term “Promoter” is used several times, for example:

“Nowadays, most ablation approaches make use of a cell type or a region-specific promoter that is coupled with a “suicide” gene, resulting in depletion of distinct cell types.”

“Unlike in the healthy CNS where microglia-targeting was achieved with the CX3CR1-promoter (see section Microglia Ablation under Healthy Conditions), all studies investigating the role of microglia in disease models use the CD11b-promoter that is also expressed in cells of myeloid origin, including both microglia and macrophages.”

“Suicide gene expression exploited for this cell population has mainly made use of the intermediate filament glial fibrillary acidic protein (GFAP) promoter that is only expressed by a subset of astrocytes in specific regions of the healthy brain (Takamiya et al., 1988; Cahoy et al., 2008).”

and so on.

I am bit confused. Which promoter is meant here? Definitely not the biological one, that is the part of DNA strand, right? Because in the first citation it is written that “region-specific promoter that is coupled with a “suicide” gene”, but both (structural) gene and promotor are part of DNA molecules/strands, how they can be coupled?

But if the “chemical” promoter is used then the situation is a bit obscure. As far as I remember for transcription not the promoter is needed but the transcription factors, right?

So, what can you tell me?

 

[BillTre]

Yes it is the biological promoter.

A promoter sequence will bind proteins (including an RNA polymerase) involved in transcribing the DNA of a gene into an mRNA.
These are usually close to the gene they affect, putting the transcriptional machinery nearby on the DNA strand. The promoter sequence tells some transcription factors where to bind the DNA. Other sequences and other factors can also be involved.
The promoter sequence can vary from gene to gene and (presumably depending on the proteins the cell is making at the time) be functional or not in different cell types.
Region specific promoters will turn on a gene in an anatomical region (like a part of the developing brain). Cell type specific promoters will turn on the gene in cells of a certain type (like microglia cells).
A suicide gene causes the cell expressing it to die.
Most likely, what they were referring to was taking out of the animal (or manufacturing) the DNA containing the promoter sequence and the suicide gene sequence and combining them in vitro (in glass, meaning in the lab not the animal), and then putting them back into the animal's genome (several times until them found a case where it worked as they wanted). Alternatively, other approaches could be used to do this, but this is most straightforward.

Thus, they get an animal that has a gene for a suicide protein controlled by the promoter for a particular cell type which will cause it to die.
If the animal can still reproduce, it could be propagated as a genetic line and bred to give more animals of a similar genetic make-up, thereby enabling a lot of related experiments.

This is kind of common in developmental biology these days. In addition, many (in theory almost any) proteins can be inserted into a genome controlled in a similar manner, for experimental purposes. Fluorescent proteins are among the most obvious examples. Here are some pictures of GFP (green fluorescent protein) in various organisms.

 

[Eagle9]

Most likely, what they were referring to was taking out of the animal (or manufacturing) the DNA containing the promoter sequence and the suicide gene sequence and combining them in vitro (in glass, meaning in the lab not the animal), and then putting them back into the animal's genome (several times until them found a case where it worked as they wanted).

So, just mixing two DNAs (one that contains promotor and second that contains suicide gene sequence) is enough for this purpose? The terminator is not needed or something else?

 

[BillTre]

So, just mixing two DNAs (one that contains promotor and second that contains suicide gene sequence) is enough for this purpose?

Well they have to be properly spliced together (linked covalently into a single DNA strand). Also the spacing and orientation have to correct.

 

[Eagle9]

Well they have to be properly spliced together (linked covalently into a single DNA strand). Also the spacing and orientation have to correct.

Ok, thanks a lot

 

[Eagle9]

Well they have to be properly spliced together (linked covalently into a single DNA strand). Also the spacing and orientation have to correct.

By the way, the Terminator is not deeded?

 

[BillTre]

By the way, the Terminator is not deeded?

You would probably want one of those too.