Below are questions and answers from previous lectures:
Wednesday, January 19, 2005:
Some excellent questions in class today (Wed 19 Jan). My guess as to the possible causes of extra Y chromosomes in humans was only partially correct. In addition to nondisjunction at meiosis II you can also get mitotic errors leading to sperm with two Y's. See paper below (note info on the frequency of the phenomenon. Keep the questions coming. I particularly enjoy these ones that I cannot answer immendiately. Thanks.....Paul
Title: The origin of the extra Y chromosome in males with a 47, XYY karyotype
Author(s): Robinson DO, Jacobs PA
Source: HUMAN MOLECULAR GENETICS 8 (12): 2205-2209 NOV 1999
Abstract: The presence of an extra Y chromosome in males is a relatively common occurrence, the 47, XYY karyotype being found in similar to 1 in 1000 male births, The error of disjunction must occur either during paternal meiosis II or as a post-zygotic mitotic error, both of which are rare events for other chromosomes, It is therefore of interest to determine when errors of Y chromosome disjunction occur, It is possible to distinguish between the different mechanisms of non-disjunction by analysing DNA polymorphisms at the distal tip of the Xp/Yp pseudoautosomal region in 47, XYY males, their parents and in some cases paternal grandparents, A cohort of 28 non-mosaic 47, XYY males was analysed. The results show that there are at least two mechanisms causing nondisjunction of the Y chromosome, In 16 of the 19 cases from which parents were available, the extra Y was generated by non-disjunction at meiosis II after a normal chiasmate meiosis I. Three cases were due to either a post-zygotic mitotic error or non-disjunction at meiosis II after a nullichiasmate meiosis I. Of the nine cases with no parental DNA available, at least four were due to meiosis II non-disjunction following a normal chiasmate meiosis I.
Monday, January 30, 2006:
Is the synthesis and attachment of primers by primase during DNA replication sequence independent or is there a set of recognition sequences at which primase begins?
First the primers in bacteria are about 10 - 12 bp long, and they are NOT random as I suggested in class. Rather, it seems that in the lagging strand, when primase comes into position to start a new Okazaki fragment, it will use the single-stranded DNA as a template to create the correct RNA primer.
Tuesday, January 31, 2006:
What guides the swapping of DNA in reciprocal translocation?
Looks like Reciprocal translocations can have many diverse causes but no evidence of any guide mechanism. Sometimes it is a function of two chromosomes both breaking at the same time and getting repaired in the incorrect alignment. The repair proteins can recognize a break point so if one chromosome breaks (and this seems to happen frequently) then the two broken ends are rejoined. If his happens to two chromosomes at the same time then the repair can cause the translocation. This process can occur at mitosis or meiosis.
Does the the initiator protein in yeast stay attached during DNA replication.
Very good question and interesting answer. In most eukaryotes it seems that the initiator proteins do indeed stay attached but NOT at the replication fork. They seem to stay attached to the origins of replication throughout the cell cycle!
Wednesday, February 4, 2004:
Why uracil and not thymine for RNA?
Thymine is created from uracil in a methylation process, which is energetically expensive. In other words, uracil is cheaper for a cell than thymine. Unfortunately cytosine becomes deaminated with significant ease to become uracil. If uracil were used in DNA, it would result in many mutations going uncorrected, as there would be no way to tell the difference between a uracil and a deaminated cytosine. By using thymine in the genetic code instead of uracil, all uracils created in such way can be removed and replaced with cytosine using the guanine base on the opposite strand of DNA as a template. The cheaper uracil is used in RNA because RNA doesn't need to last a long time anyway.
Are hairpin loops universal?
No -They occur in some prokaryotes (other prokaryotes have rho factor sites). Also, they occur in some (not all) eukaryotes.
Do the hairpin loops get cleaned up off of mRNA?
Mine don't.
What are the 5' cap enzymes?
Capping enzymes:
1) guanylyl transferase - Adds the 5' (backwards G) [includes the triphosphate bridge)
2) Methylation of terminal G by guanine-7-methyl transferase
3) the 2' (-O) on the pentose of the second base is methylated by 2'-O-methyl transferase
4) In some euk. mRNAs methylation occurs to the second base only if it is an adenine (N6 position on adenine)
4)Sometime also to the third base. 3 and 4 are methylated by other methyltransferases
Thursday, February 5, 2004:
Is RNA editing found in prokaryotes? Do they perform the U to C or C to U editing?
RNA editing not known in prokaryotes nor in yeast
Can side loops in a tRNA function as the anti-codon?
Update: I am still unable to find published evidence of a tRNA functioning with an anticodon in a side loop. However, there are functional organellar (chloroplast and mitochondrial) tRNAs that occassionally (rarely) lack the D-loop (they are called "amputated tRNAs")
Is the modified bases, such as Inosine, used only for the third codon in a tRNA or can they be used for the first or second codon as well?
Probably yes. Base modification is common throughout trNA molecules. However, I am still looking for documentation of modification of the anticodon outside the wobble position. I am guessing that the 3' base is modified when GUG is a start codon (see below).
How is GUG recognized?
By tRNA-fMET for sure. No literature detected but I speculate that The 3' position of the anticodon is modified to pair with either purine.
How are the tRNA molecules released from the ribosomal subunits?
Once the peptide bond forms between the amino acid at the P site and that at the A site, the reaction that creates that covalent bond first has to break the bond holding the amino acid to the tRNA (at the P site). Thus, the peptidyl transferase is involved in release the tRNA.
Tuesday, February 7, 2006:
What is the origin of introns? Why are they there?
Several hypotheses:
1. Relicts of transposable elements
2. Hypothesis 1 suggests that too much of the cellular machinery is invested for this to be a good explanation so an alternative is that introns are there for control and regulation of gene expression.
No clear consensus at the moment. Abstract below on retrotransposon theory and another attached. Along with hypotheses on the selective forces involved there are compewting hypotheses as to whether introns were acquired early or late in eukaryotic evolution. Have they been lost in prokaryotes?
Introns as relict retrotransposons: implications for the evolutionary origin of eukaryotic mRNA splicing mechanisms.
Hickey DA, Benkel B.
J Theor Biol. 1986 Aug 7;121(3):283-91.
A model is presented for the evolutionary origin of intron sequences within eukaryotic protein-coding genes. We propose that introns are the vestiges of transposable elements and, specifically, that they represent a novel class of retrovirus-like transposons. The attraction of the retrotransposon model is that it gives the RNA splicing mechanism a central role in the evolution of introns. There is a growing body of evidence to suggest that several aspects of splicing are intron-encoded. Consequently, it is reasonable to look for evolutionary explanations of the splicing mechanism in the context of the evolution of the intron sequences themselves. According to this model the ancestral intron genomes were replicated into RNA copies simply because of their insertion within transcriptionally active regions of the host genome. Splicing was necessary not only to minimize their negative effects on host gene expression, but also, and perhaps more importantly, to generate new copies of the intron genome free of flanking exon sequences. These spliced intron copies were then available for reverse transcription and reinsertion elsewhere in the genome. Thus, splicing can be seen as an essential step in the intron replication cycle. Most modern introns have probably lost the majority of their original genetic content and may be considered as degenerate evolutionary relicts. An exception to this degeneracy is the set of splicing signals which must be retained because of its continued importance to host cell survival.(ABSTRACT TRUNCATED AT 250 WORDS)
PMID: 3025526 [PubMed - indexed for MEDLINE]
How can you begin in vitro translation without a start codon?
In short, you can't start translation without a start codon. In Nirenberg's experiments to decipher the genetic code, their in vitro translation reactions included purified RNA from E. coli and the the system was already operating (initiated). All they did was add the poly-U RNA and recorded increases of radioactive proteins. They found the labeled poly-Phe increased in response.
Wednesday, February 8, 2006:
In prokaryotes, which binds to the mRNA first, the fMet tRNA or the
small sub-unit of the ribosome?
The SSU of the ribosome attaches first.
Thursday, March 2, 2006:
How do Barr bodies replicate during normal cell division and what
controls ensure that the same copy of the chromosome in the parent cell
ends up being the Barr bodies in the two daughter cells?
Answer: There is some evidence of methylation so that the Barr body
gets replicated like all other X-chromosomes but then the methylation
pattern guides reheterochromatinization going into the next interphase.
Wednesday, March 3, 2004:
Is it more common to have a homozygote or heterozygote for inversions?
Only really apparent that you have an inversion when you examine meiosis in a heterozygote. Some inversion heterozygotes are quite stable. Difficult to say in general which is most common
Which came first- the DNA sequence or enzyme pathway for the gene associated with chronic myelogenous leukemia?
Still researching this.
Are microsatellites remnants of TEs?
Cannot find any evidence for this or any record of such a suggestion. Microsatellites appear to evolve from errors in replication.
Thursday, March 4, 2004:
At what phase of the cell cycle do the transposons move?
No literature found on this topic. I would think they could move throughout the entire cell cycle.
Are SINES and LINES remnants of retroposons?
I believe so since SINES and LINES are called retroposons in the literature.
How do you use sister chromatids as templates?
No details found in the literature. It might not be known at this time. The literature has discussed why sister chromatids are used as templates and how they separate.
Kadyk, L.C. and Hartwell, L.H. 1992. Sister chromatids are preferred over homologs as substrates for recombinational repair in Saccharomyces cerevisiae . Genetics , 132: 387-402.
Matioli, G.T. 1991. On the handedness of the scaffolds of sister chromatids. Medical Hypotheses , 35: 52-58.
Can nondisjunctions (mosaics) cause cancer?
It is thought to have some role in causing cancer, although as far as we can tell from the literature, it is unknown at this time.
Monday, March 15, 2004:
How did plasmids arise?
Still researching how plasmids arose. Found the suggestion by French scientists that plasmids could arise by the uptake (transformation) by soil bacteria of random peices of "environmental" DNA (partially degraded from plants, animals, fungi). Such transformation could occur as a result of lightening which could act as electroporation, making cells competant. Most bits of DNA taken up likethis would be lost but if occassionally a peice contained an origin of replication AND became circular then it could continue to replicate. Perhaps there have been multiple origins in this way.
I have also come across the suggestion that some bacteriophages may have evolved from plasmids.
Tuesday, March 16, 2004:
What factors influence a shift from lytic to lysogenic, and vice versa, in bacteriophages?
Results from a study suggested that the relative amounts of the two proteins ORF4 and ORF5 determine the decision between lytic or lysogenic life cycle after phage infection and that a protein complex consisting of ORF4 and ORF5 may constitute a new type of genetic switch in bacteriophages.
When a host bacterial cell is naturally transformed and they take up a gene that is not identical to their own genome, does the bacteria have mechanisms for repairing the mis-matched strands of DNA? Basically, why does the new strand of DNA get incorporated into the host cells genome? Aren't there mechanisms to prevent this, such as methylation of the host cells DNA?
I am still working on the information from bacterial cells.
In mammalian cells, foreign DNA is extensively de novo methylated in specific patterns when it is inserted into the genome. This methylation is part of the mammalian host cell defense mechanism. So the genome can contain foreign DNA that is methylated. The methylation is random, so the likelihood that the gene you are trying to transform the cell with is methylated is relatively low.
Doerfler, W. 1996. A new concept in (adenoviral) oncogenesis: integration of foreign DNA and its consequences. Biochimica et Biophysica Acta , 1288: F79-F99.
When an Hfr cell is transferring plasmid and host cell DNA to an F- cell, can that host cell plus plasmid DNA become a new plasmid during conjugation?
No studies have been done on this. I don't really see why it could not become a new plasmid, especially if it is not incorporated into the cell's genomic DNA.
Wednesday, March 17, 2004:
Today's question dealt with the number of introns in mitochondria of animals versus plants.
It depends on the organism you are looking at. It seems that most mitochondria of plants have fewer introns than mitochondria of animals. But the mitochondria in plants and animals both come from the same ancestor.
Is there a relationship between mutant mtDNA and brain development? Basically, why is the brain the first organ affected when there are mutant mitochondria?
A study done on mice revealed that copper is essential for the activity of certain neurotransmitter systems. Mitochondria contain enzymes that require copper to function normally. This study found that the progressive decline in brain metabolism might be related to the abnormal development of the copper-requiring mitochondrial enzymes. The study also indicated that the brain requires a large amount of glucose and goes through oxidative metabolism during development. This is probably why the brain is the first organ affected with mutant mitochondria.
Another study with mice has shown that mutations in the mitochondrial CAP gene (a mitochondrial ribosome inhibitor-chloramphenicol) resulted in increased mitochondrial production of reactive oxygen species throughout the development of the early eye altering the lens crystalline proteins. This could explain why cataracts form in the elderly.
Peterson, C.E. and Goldman, J.E. 1986. In vitro acetylcholine synthesis and oxidative metabolism during development of normal and brindled mouse brain. Developmental Brain Research , 29: 153-159.
Sligh, J.E. et. al. 2000. Maternal germ-line transmission of mutant mtDNAs from embryonic stem cell-derived chimeric mice. P.N.A.S. , 97 (26): 14461-14466.
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