Any microbiologists? DNA question

[svero]

So I'm reading the book DNA by Watson. In it he describes a process where bacteria gain protection/resistance by sharing plasmids and how elements of that process are used to insert plasmids from other animals.. say a strand of toad DNA into bacteria so they can be copied during regular bacterial cell division. It brings up a few questions that aren't addressed in the book. If someone could fill me in I'd appreciate it.

1) Do the plasmids themselves code in the protection? That is to say... Are the plasmids somehow incorporated into the bacteria's DNA after splitting or does the nucleus contain both the bacterias dna and the plasmids which are then passed on through subsequent generations.

2) This is related to the first question.. but normally does the cell nucleus contain just 1 strand of dna with all the genes or is it usually several strands with different strands having different genes? How about in humans. I get the impression that the human genome is 1 long strand of dna.. are there also extra bits in there that get copied?

3) when a virus that uses reverse transcriptase to add dna to a cell does it's thing... does the viruses dna become part of the cells dna? Say HIV for instance... It infects a cell, and then new dna is introduced. If I've got it right the cell now is essentially converted into an hiv factory. Is the cells initial dna now modified to incorporate the virus dna or is it two separate strands in the nucleus. If the infected cell devides (assuming it can still devide after being infected - and assuming t helper cells ever do devide - both things im unsure of) I assume it would also copy the viruses dna?

Anyway.. obviously I'm quite ignorant... but if someone can try to answer those questions I'd appreciate it.

 

[Prester John]

1) Plasmids are independently replicating genetic elements. Often circular, but they can be linear. They carry genes that code for their own replication. The best studied are those that confer resistance to antibiotics. I like to think of plasmids as power ups (well maybe not quite but its an analogy).

2) Theres lots of DNA in even a bacterial cell. eg E.coli has about 1mm of DNA (about 1000 times its length). Plasmids show that this can be in several distinct elements, but then most plasmids probably aren't neccessary for bacterial survival. A bacterium doesn't have a nucleus like a human does. Additionally our genetic material is split into chromosomes.

3) Integrated into the cells DNA, thus if the cell was capable of dividing then the viral DNA would get copied as well.

 

[svero]

Just to expand on your answer in question 2 - In our DNA then each chromosome is a separate strand of DNA? Are these strands looping or linear? or some of both?

 

[Prester John]

http://www.sci.sdsu.edu/~smaloy/MicrobialGenetics/topics/chroms-genes-prots/chromosomes.html

http://en.wikipedia.org/wiki/Chromosome

Should answer the questions (ie yes and linear)

 

[American]

Any microbiologists? DNA question

Not to nit-pick, but microbiology is usually considered on a microscopic scale (ie, you may see bacteria or whatever using a microscope).

When you talk about DNA questions, you're likely on a molecular scale, and hence it's a molecular biology question.

There is, naturally, a great deal of overlap, so in many cases either term may be accurate (particularly as you refer to plasmids, which are an exclusive tool of microbiology).

 

[Jon_in_london]

A plasmid is BY DEFINITION not essential to a bacteria's survival under normal conditions (ie no antibiotics present etc...) If it IS essential then it is classed as a chromosome rather than plasmid.

Fascinating things plasmids...... almost a virus... but not quite.

 

[Jon_in_london]

3) when a virus that uses reverse transcriptase to add dna to a cell does it's thing... does the viruses dna become part of the cells dna? Say HIV for instance... It infects a cell, and then new dna is introduced. If I've got it right the cell now is essentially converted into an hiv factory. Is the cells initial dna now modified to incorporate the virus dna or is it two separate strands in the nucleus. If the infected cell devides (assuming it can still devide after being infected - and assuming t helper cells ever do devide - both things im unsure of) I assume it would also copy the viruses dna?

Sometimes, some viruses will integrate their genome into the host's DNA (as HIV can do) and then lurk around for a while. Incidentally, a significant proportion of the human genome is of retroviral origin. However others do not- they simply order the cell to make loads of copies of virus until the cell pops..

Yes, T cells can and do divide, in fact they can do so in spectacular fashion when they need to. Yes, the viral DNA will be copied but only in half its progeny if there is only one viral copy in its DNA.

 

[svero]

Re: Re: Any microbiologists? DNA question

Sometimes, some viruses will integrate their genome into the host's DNA (as HIV can do) and then lurk around for a while. Incidentally, a significant proportion of the human genome is of retroviral origin.

Are you saying a lot of the so called "Junk" Dna is from retroviral additions? or that much of our actual coding dna was a sort of forced mutation we devloped as a species?

However others do not- they simply order the cell to make loads of copies of virus until the cell pops..

Well I hadn't intended this thread to sink into a discussion of HIV but you bring up a lot of questions that have floated around in my head on this so here goes... Is it one of the difficulties with "curing" HIV that HIV is able to lurk around in cells that look normal to the body? Like.. why is it that antibiotics can kill off a throat infection but anti-retrovirals can't totally kill off HIV? Is it that HIV is better at mutating and resisting or is it that it's integrated itself into our DNA somewhere and the action of the anti-retroviral is no longer a killer (say messing reverse transcriptase up..). Too late it's already in the dna. I've never understood how HIV lurks and recovers after a patient comes off anti-retrovirals.

Yes, T cells can and do divide, in fact they can do so in spectacular fashion when they need to. Yes, the viral DNA will be copied but only in half its progeny if there is only one viral copy in its DNA.

I don't understand what you mean by half? Can you explain this more clearly? Wouldn't both of the devided cells contain the same DNA after a split? and therefor both be HIV factories? And if the t helpers just have extra dna then why are they no longer useful at fighting off other diseases. Does the integration of the virus completely destroy their capabilty in the immune system or is it that the t-helpers die shortly after infection? Where do the t-helper cells normally originate from? Are they created in the thymus? That is to say... suppose someone is infected with HIV... and some of the t cells have the dna already encoded. Is it hypothetically possible to rid the body of all those cells and have it create new uninfected t-cells? I would assume that's correct since people on retrovirals will see their t cells counts return to normal levels of the drugs work.

Hehehe... lots of questions... Bet you're sorry you stuck your nose in this thread now!

 

[svero]

A plasmid is BY DEFINITION not essential to a bacteria's survival under normal conditions (ie no antibiotics present etc...) If it IS essential then it is classed as a chromosome rather than plasmid.

Fascinating things plasmids...... almost a virus... but not quite.

Thanks. I was going to ask that. Searches on google for "difference chromosome plasmid" yielded too many pages to sort through. It was a natural question.

 

[Jon_in_london]

Thanks. I was going to ask that. Searches on google for "difference chromosome plasmid" yielded too many pages to sort through. It was a natural question.

You can also get 'episomes' which are just bits of DNA that have been chopped out of the chromosome and circularised. However, an episome is only a plasmid if it has an replicating mechanism and usually some method to ensure it isnt lost when the host cell divides.

 

[Jon_in_london]

Re: Re: Re: Any microbiologists? DNA question

Are you saying a lot of the so called "Junk" Dna is from retroviral additions? or that much of our actual coding dna was a sort of forced mutation we devloped as a species?

Im not sure about whether or not its 'junk' or coding or maybe a bit of both. Probably the latter.

Well I hadn't intended this thread to sink into a discussion of HIV but you bring up a lot of questions that have floated around in my head on this so here goes... Is it one of the difficulties with "curing" HIV that HIV is able to lurk around in cells that look normal to the body? Like.. why is it that antibiotics can kill off a throat infection but anti-retrovirals can't totally kill off HIV? Is it that HIV is better at mutating and resisting or is it that it's integrated itself into our DNA somewhere and the action of the anti-retroviral is no longer a killer (say messing reverse transcriptase up..). Too late it's already in the dna. I've never understood how HIV lurks and recovers after a patient comes off anti-retrovirals.

Yes, HIV is an expert 'lurker'. Its mode of action is highly specialised to allow it to lurk around our lympathic system- which is kind like Al-Qaeda hiding out in CIA Headquarters! But in order to do so, it causes great damage to the lymphatic system which is why AIDS develops. Im not too sure about why anti-retrovirals become ineffective but I assume its due to selective pressure selecting in the resistant ones (?) HIV can survive a blitz of retrovirals because it can lie dormant and not be affected by the drug

I don't understand what you mean by half? Can you explain this more clearly? Wouldn't both of the devided cells contain the same DNA after a split? and therefor both be HIV factories? And if the t helpers just have extra dna then why are they no longer useful at fighting off other diseases. Does the integration of the virus completely destroy their capabilty in the immune system or is it that the t-helpers die shortly after infection? Where do the t-helper cells normally originate from? Are they created in the thymus? That is to say... suppose someone is infected with HIV... and some of the t cells have the dna already encoded. Is it hypothetically possible to rid the body of all those cells and have it create new uninfected t-cells? I would assume that's correct since people on retrovirals will see their t cells counts return to normal levels of the drugs work.

What do I mean by half? Brush up on Mendelian genetics- we have two copies of our DNA- half goes into one progeny cell and half into the other.

Each T-cell is specifically geared to fight a single antigen or disease if you like......... eeeesh!!! Big Questions!!! I have to go to work but I'll try and answer this lot later.


Hehehe... lots of questions... Bet you're sorry you stuck your nose in this thread now!

Thats OK!

 

[Capsid]

Like.. why is it that antibiotics can kill off a throat infection but anti-retrovirals can't totally kill off HIV

Antibiotics (usual cause of a throat infection) work on bacteria NOT viruses because the bacteria do not integrate into the host's genome like HIV and other integrating viruses do. So the antibiotic can kill the free living bacteria but not viruses which may persist merely as a piece of DNA (a provirus).

So treatment with retrovirals will suppress or inhibit the replicative process of viruses but not rid the body of the proviral DNA. Stopping treatment allows the replicative process to function again and thereby produce viral particles.

I don't understand what you mean by half?

The proviral DNA may integrate into just one of the chromosome pairs so when the cell divides the provirus will only be transferred to one daughter cell.

The T cells can continue to fight other infections after integration by HIV and also the CD8 T cells can control HIV for some time. But eventually the CD4 cells are destroyed by HIV and their numbers decline to such a low level that their role in the immune system is not sufficient to ward off opportunistic infections.

T cells originate from the bone marrow and mature in the thymus.

Retrovirals don't work by killing virus, only the immune system can do that through CD8 cells or by antibody neutralisation. But as we've seen the immune system is severely compromised by HIV (it's not called immunodeficiency virus for nothing). If you want drugs to kill viruses then they will have to kill the cells in which the provirus resides, this would be very difficult to achieve.

 

[svero]

So treatment with retrovirals will suppress or inhibit the replicative process of viruses but not rid the body of the proviral DNA. Stopping treatment allows the replicative process to function again and thereby produce viral particles.

Ugh.. no wonder its so hard to cure. Sounds impossible! The resistance and inability of anti retrovirals to kill the disease off completely makes perfect sense now though. We need a retroviral dna cleaning virus! Which will of course mutate into a superflu.

The proviral DNA may integrate into just one of the chromosome pairs so when the cell divides the provirus will only be transferred to one daughter cell.

Hmm strange. I would have expected that the extra bits had to be paired off to fit into the structure nicely.

The T cells can continue to fight other infections after integration by HIV and also the CD8 T cells can control HIV for some time. But eventually the CD4 cells are destroyed by HIV and their numbers decline to such a low level that their role in the immune system is not sufficient to ward off opportunistic infections.

T cells originate from the bone marrow and mature in the thymus.

Retrovirals don't work by killing virus, only the immune system can do that through CD8 cells or by antibody neutralisation. But as we've seen the immune system is severely compromised by HIV (it's not called immunodeficiency virus for nothing). If you want drugs to kill viruses then they will have to kill the cells in which the provirus resides, this would be very difficult to achieve.

Right because the HIV infected cells look identical to the non-infected cells if im not mistaken.

Just musing here.. I assume this has all been thought of before, but maybe what's needed is a technology that can somehow modify the cells to tag themselves when they're infected, or which tags infected cells as they're infected. Suppose somone was on an anti-retroviral drug and tagging drug regimin is it possible that eventually all the infected cells would be tagged if we could assume that the ones that were infected prior to the drug regimin die off? Then one could hypothetically wait some necessary safe time period and take a tag killing drug that would target and kill all tagged cells. Or alternately they could be tagged in such a way that the immune system recognizes them as intruders and takes them out. Im assuming that not so many t cells are tagged that the tag killing action itself would immediately decimate the immune system.

 

[Capsid]

Virus infected cells are tagged by presenting viral proteins on class I molecules on their cell surface. So they are different (subtly) from normal cells. The viral proteins are recognised by CD8 cells which can kill the cell directly and thereby the virus. So the immune system can exquisitely identify viruses and normally it is able to control viral infections very well except where the immune system is compromised in some way. Chronic hepatitis B and hepatitis C are examples of chronic viral infections where the immune response against the virus is weak allowing the virus to persist.

Technologies that are close to what you suggest have involved anti-cancer antibodies carrying radioactivity which can kill the cancer cell (not sure how successful or widely used this is though). This hasn't been translated to HIV, the problem being that it would be difficut to hit all the virus infected cells and then killing the very cells (CD4 T helper cells) that the immune system needs to control the virus is not good and would probably accelerate progression to AIDS.

The preferred route is to develop a vaccine (therapeutic and prophylactic).

 

[Shane Costello]

I don't like letting DNA threads slip by without making some sort of contribution, so apologies if the following seems trite and nitpicky.

The term junk DNA is fast falling from favour. It erroneously implies that DNA that is neither a gene nor involved in gene transcription is of no value. Instead it appears that these DNA sequences can play vital roles in areas such as RNA and genomic stability. Non-coding DNA is the preferred term. Bacterial genomes have much less non-coding DNA than eukaryotic genomes. An important point to note relevant to plasmid genetics is the fact that bacterial genes do not contain promoter elements. This is an important consideration when it comes to using plasmids to insert eukaryotic genes into bacteria, since eukaryotic genes require promoter elements to be expressed. It makes the gnetic transformation of bacteria with, for instance, a toad gene that bit trickier.

 

[Jon_in_london]

I don't like letting DNA threads slip by without making some sort of contribution, so apologies if the following seems trite and nitpicky.

The term junk DNA is fast falling from favour. It erroneously implies that DNA that is neither a gene nor involved in gene transcription is of no value. Instead it appears that these DNA sequences can play vital roles in areas such as RNA and genomic stability. Non-coding DNA is the preferred term. Bacterial genomes have much less non-coding DNA than eukaryotic genomes. An important point to note relevant to plasmid genetics is the fact that bacterial genes do not contain promoter elements. This is an important consideration when it comes to using plasmids to insert eukaryotic genes into bacteria, since eukaryotic genes require promoter elements to be expressed. It makes the gnetic transformation of bacteria with, for instance, a toad gene that bit trickier.

Im sorry Shane but Bacterial genes most certainly DO have promoters (LacZ?)! However, a prokaryotic promoter wont necesarily work in a eukaryote. Indeed, promoter activity levels vary greatly even between one higher plant and another.

 

[Jon_in_london]

Virus infected cells are tagged by presenting viral proteins on class I molecules on their cell surface. So they are different (subtly) from normal cells. The viral proteins are recognised by CD8 cells which can kill the cell directly and thereby the virus. So the immune system can exquisitely identify viruses and normally it is able to control viral infections very well except where the immune system is compromised in some way. Chronic hepatitis B and hepatitis C are examples of chronic viral infections where the immune response against the virus is weak allowing the virus to persist.

Actually, HIV issues a recall signal (Vpr) so that infected CD4+ withdraw their MHC and therefore become 'invisible' to the CD8+. This wouldnt really cause any problems except HIV infected cells also secrete Nef- which is quite similar to bee-sting venom. This wreaks wanton destruction on the lymphatic system and eventually leads to AIDS. HIV does NOT set out to kill its host cells, it is in fact very clever at ensuring its host's survival- to the detriment of the immune system as a whole.

Technologies that are close to what you suggest have involved anti-cancer antibodies carrying radioactivity which can kill the cancer cell (not sure how successful or widely used this is though). This hasn't been translated to HIV, the problem being that it would be difficut to hit all the virus infected cells and then killing the very cells (CD4 T helper cells) that the immune system needs to control the virus is not good and would probably accelerate progression to AIDS.

You dont really need to make the anibodies radioactive. I know of at least one trial in which cancer patients were 'vaccinated' with their own tumour material. The hope being that the immune system would now recognize the cancer as non-self and attack it. As far as I remember, the study had some success with remission times being generally better for the vaccinated sample.

The preferred route is to develop a vaccine (therapeutic and prophylactic).

Another option would be to use a gene therapy that would insert a gene that would be lethal only in the prescence of HIV. I dont know if any studies have been dome around this.

 

[svero]

Actually, HIV issues a recall signal (Vpr) so that infected CD4+ withdraw their MHC and therefore become 'invisible' to the CD8+. This wouldnt really cause any problems except HIV infected cells also secrete Nef- which is quite similar to bee-sting venom. This wreaks wanton destruction on the lymphatic system and eventually leads to AIDS. HIV does NOT set out to kill its host cells, it is in fact very clever at ensuring its host's survival- to the detriment of the immune system as a whole.

So it's the secreted Nef- that actually causes the destruction of CD4 cells and not the genetic transformation in the cell itself?

 

[Jon_in_london]

So it's the secreted Nef- that actually causes the destruction of CD4 cells and not the genetic transformation in the cell itself?

Nef is the thing that causes AIDS. Without Nef, HIV would be relatively harmless parasite.

Its important to recognize though, that HIV doesnt want to kill CD4s- those cells are its home. It does want to kill the rest of the immune system for its own self-defence.

 

[svero]

Nef is the thing that causes AIDS. Without Nef, HIV would be relatively harmless parasite.

Its important to recognize though, that HIV doesnt want to kill CD4s- those cells are its home. It does want to kill the rest of the immune system for its own self-defence.

But I was under the impression that it was the CD4 cells that you lost when infected. And that once their count went below a certain amount the immune system was no longer able to protect you against other diseases. If HIV doesn't want to kill it's home why is it principly the cd4 counts that are looked at in patients and reduced? This seems counter intuitive to what I've previously read.