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ariariari ariariari
wrote...
Posts: 3
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9 years ago
So this is the toughest question I got from my cell bio class. I've spent last two days trying to solve this and I am going no where.

To examine the hierarchy of signal sequence targeting in a cell, you construct a variety of proteins containing different combinations of signal sequences. Based on your extensive knowledge of protein targeting, predict the location of each of these fusion proteins listed below. Provide the logic for your predictions.

(A) ER signal sequence and NLS
I know how both of them work very well. I first thought it would end up in ER cuz I thought protein with NLS will be brought back out to cytosol and but will be stuck in ER. But of course, I just realized that it doesn't have NES so once in nucleus, the protein will be stuck in nucleus.

(B) mitochondrial signal sequence and KDEL sequence
Assuming the protein starts in the cytosol, KDEL will not have any effect since the protein is not in ER or Golgi network - will have to be brought into ER first but doesn't have ER signal sequence! So the protein will just end up in mitochondria or on its membrane depending on the type of signal sequence.

Any suggestion about ER signal sequence vs NLS?
Or anyone that disagree with mito vs KDEL sequence?
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wrote...
9 years ago Edited: 9 years ago, soniasunny
for part A...hope this helps
An example of such a targeting sequence is the so-called nuclear localization signal (NLS), which consists of two short stretches of basic amino acids divided by a 10–11 amino acid spacer region. This sequence of amino acids allows a protein possessing it to bind to nuclear localization receptors found in the nucleus . Once a protein containing an NLS signal binds to a nuclear receptor it is no longer able to freely diffuse and becomes "localized" to the nucleus.

The interest that exists in NLSs is not only for investigating endogenous purposes, but also for the engineering of other proteins to be imported into the nucleus. The signal is small (4-8 amino acid residues, or 12-24 base pairs), therefore it is easily imbedded in the sequences of such reporter genes as eGFP and beta-galactosidase. Expression of the protein in the nucleus means that it is due to your gene being expressed in a cell, which is of some concern with proteins targeted to the cytoplasm. These proteins that lack NLS targeting can be translocated across intracellular space and be expressed in other cells. This means that the protein that your gene is creating is giving false positives, in a mechanism known as pseudotransduction. The NLS targeting of a protein traps that protein within one cell, and does not allow for this phenomenon.
Please read details at
http://everything2.com/title/Nuclear+Localization+Signal

Post Merge: 9 years ago

for part B

how does the protein end up in mitochondria?
the protein itself is created within the ER and reaches the golgi apparatus from where the signal is added the KDEL and then targeted to the site, in this case it the KDEL which will automatically drives it to  ER lumen. this is because KDEL sequence is  found on proteins that are supposed to reside in the ER lumen after synthesis.  If by chance a protein with the KDEL sequence is packaged into vesicles released from the ER, the golgi will send it back to the ER.
for the protein to be sorted and packaged to the mitochondria we need a different protein tag.
Mitochondrial matrix
The signal peptide that directs to the mitochondrial matrix has a sequence consisting of an alternating pattern with a few hydrophobic amino acids and a few plus-charged amino acids form. It is usually called the mitochondrial targeting signal (MTS).

hope this makes sense....
ariariari Author
wrote...
9 years ago
for part A...hope this helps
An example of such a targeting sequence is the so-called nuclear localization signal (NLS), which consists of two short stretches of basic amino acids divided by a 10–11 amino acid spacer region. This sequence of amino acids allows a protein possessing it to bind to nuclear localization receptors found in the nucleus . Once a protein containing an NLS signal binds to a nuclear receptor it is no longer able to freely diffuse and becomes "localized" to the nucleus.

The interest that exists in NLSs is not only for investigating endogenous purposes, but also for the engineering of other proteins to be imported into the nucleus. The signal is small (4-8 amino acid residues, or 12-24 base pairs), therefore it is easily imbedded in the sequences of such reporter genes as eGFP and beta-galactosidase. Expression of the protein in the nucleus means that it is due to your gene being expressed in a cell, which is of some concern with proteins targeted to the cytoplasm. These proteins that lack NLS targeting can be translocated across intracellular space and be expressed in other cells. This means that the protein that your gene is creating is giving false positives, in a mechanism known as pseudotransduction. The NLS targeting of a protein traps that protein within one cell, and does not allow for this phenomenon.
Please read details at
http://everything2.com/title/Nuclear+Localization+Signal

Post Merge: 9 years ago

for part B

how does the protein end up in mitochondria?
the protein itself is created within the ER and reaches the golgi apparatus from where the signal is added the KDEL and then targeted to the site, in this case it the KDEL which will automatically drives it to  ER lumen. this is because KDEL sequence is  found on proteins that are supposed to reside in the ER lumen after synthesis.  If by chance a protein with the KDEL sequence is packaged into vesicles released from the ER, the golgi will send it back to the ER.
for the protein to be sorted and packaged to the mitochondria we need a different protein tag.
Mitochondrial matrix
The signal peptide that directs to the mitochondrial matrix has a sequence consisting of an alternating pattern with a few hydrophobic amino acids and a few plus-charged amino acids form. It is usually called the mitochondrial targeting signal (MTS).

hope this makes sense....

I'm not sure if this is how I reply to your reply.

(A) I still don't know whether the nuclear-encoded mRNAs trnaslated on cytosolic ribosomes will be directed to nucleus ER. I understand that proteins with NLS will only work for that cell (and in this case will be prolly get stuck in nucleus since it has no NES) but not sure if there's a mechanism that prevents either NLS or ER signal from happening.

(B) The protein starts in the cytosol as a complex of mRNA + ribosomes + polypeptide. It would be stuck in ER if the protein does start within the ER just like you said but I think the question is asking where the protein would go if it started in cytosol. This protein has no ER signal so won't be able to enter ER in first place, so synthesis of this protein will be completed on free ribosomes in cytosol - then get directed to mitochondria.
wrote...
9 years ago
The synthesis of all proteins begins in the cytosol (the nonparticulate part of the cytoplasm) .

However, proteins that will finish their synthesis on the RER have a sequence of amino acids called the signal sequence because this series of amino acids acts as a signal. Signal sequences are specific sequences of amino acids of a newly forming (nascent) polypeptide chain.

Proteins that will be completely synthesized in the cytosol do not have a signal sequence.


http://academic.brooklyn.cuny.edu/biology/bio4fv/page/er_funct.htm

check the above link
ariariari Author
wrote...
9 years ago
The synthesis of all proteins begins in the cytosol (the nonparticulate part of the cytoplasm) .

However, proteins that will finish their synthesis on the RER have a sequence of amino acids called the signal sequence because this series of amino acids acts as a signal. Signal sequences are specific sequences of amino acids of a newly forming (nascent) polypeptide chain.

Proteins that will be completely synthesized in the cytosol do not have a signal sequence.


http://academic.brooklyn.cuny.edu/biology/bio4fv/page/er_funct.htm

check the above link


Thanks for the info but I think you're misreading the question. The quesiton is asking what if that protein has both ER signal sequence and NLS..
wrote...
9 years ago
opps...


if so then then even I am confused as why should a protein have both the signals...
it must be withe either one of the signal to get directed to the particular organelle....
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