| M W F; 10:10 - 11:00 pm |
Molecular Biology |
Douglas W. Smith |
| York 2722 |
BIMM 100 |
5254 Muir Biology Building |
| Fall, 2000 |
x42620; dsmith@ucsd.edu |
| BIMM100 | Syllabus
| Sections / Off Hrs | Grading
Policy | DNASYSTEM
|
| Lectures | Journal
Articles | Study Qs | Lab
Techniques | Exams |
What types of RNA molecules are made by eukaryotic RNA polymerases?
Which RNA polymerases catalyze synthesis of which RNA types?
Where are each of the RNA polymerase types found in the eukaryotic cell?
What is the Nucleolus? the Nucleoplasm?
How does the structure of a eukaryotic CoreRNAPol compare with the E. coli CoreRNApol?
What are RNA Pol "basal" factors?
What are RNA Pol "Transcription factors"?
What E. coli protein is comparable to eukaryotic Transcription Factors?
What is the nature of the eukaryotic Promoter?
Why is it divided into a Core Promoter and Enhancer sites?
What is the Core Promoter and where is it located relative to the Transcription start site?
What are Enhancer sites? Are they upstream or downstream of the Core Promoter?
What is the basic structure of the genes transcribed by RNA Pol I?
What is the structure of the Pol I promoter region?
What do the terms UCE, UBF1, and SL1 mean?
Which of the transcription factors is most similar to the E. coli sigma factor?
What is TBP and what is its function? What is the structure of TBP?
What are the "Basal Apparatus Proteins" of RNA Pol II? What other proteins are there? What is the function of these other proteins?
What promoter sites do the Basal Apparatus Protein need for their function?
What is the function of TFIIA? TFIIB? TFIID? TFIIE? TFIIF? TFIIH? TFIIH?
Is TBP involved with RNA Pol II? If so, how?
What is the "Initiator" (Inr) site and what is its function?
Are all interactions Protein-DNA interactions? If not, what others are important?
Which RNA Pol II basal proteins have DNA sequence specificity?
Do all RNA Pol II promoters have a TATA box? What happens if one doesn't?
What is the basic structure of each of the promoters used by RNA Pol III and what types of RNA are transcribed from each of these promoters?
What are the functions of transcription factors TFIIIA and TFIIIC?
Why is TFIIIB thought to play a role similar to E. coli sigma factor?
What site and what molecule provide specificity for where transcription begins at each of these 3 types of promoters?
What does PSE mean? What does DSE mean?
What is a "TATA box" and what is its function? Do all promoters have TATA boxes? Why or why not? What provides the function of the TATA box when it isn't present?
What is a PreInitiation Complex? What fundamental difference in prokaryotic and eukaryotic transcription initiation does this concept signify?
What processing events occur with prokaryotic mRNA?
What is the usual half-life of prokaryotic mRNA? of eukaryotic mRNA?
What does "half-life of mRNA" mean?
What is the fate to the prokaryotic mRNA? How does this occur enzymatically?
Prokaryotic mRNA is often polycistronic. What does this mean?
What does "coordinate expression" mean? How is this related to the above statement?
What is a cistron? a recon? a muton?
Are there more, or less, translational start sites than transcriptional start sites? Think about this in terms of a polycistronic mRNA.
It is said: Transcription and translation are coupled in bacteria. What does this mean? How is this related to the fate of prokaryotic mRNA? How are the rates of transcription and translation consistent with this statement? What are these rates?
How are polysomes related to this issue of coupling? What are polysomes?
How can mRNA secondary structure be used as a control mechanism for gene expression? Why is polarity often the result of mutations in one of the expressed cistrons? Mutations in which cistron? What is polarity?
How does the above control mechanism differ from control via ribosomal frameshift? What is a frameshift?
Eukaryotic mRNA is usually monocistronic. What does this mean? Why is this the case for eukaryotes, whereas polycistronic mRNA is often found in prokaryotes?
Transcription and translation are not couples in eukaryotes. Why is this the case?
Name four post-transcriptional modification events which can occur on eukaryotic mRNA. Which of these ALWAYS occur? Which USUALLY occur?
What eukaryotic mRNA modification event involves poly(A)? What is poly(A)? What is Poly(A)-Binding Protein? How does it work and what does it do?
How is the poly(A) modification used to purify eukaryotic mRNA? How does oligo(T) differ from poly(A)? What RNA does this purification step separate eukaryotic mRNA from? Why is this RNA otherwise a major problem?
What common eukaryotic proteins are usually poly(A)-free?
What is the first step in capping eukaryotic mRNA? Where is the Cap? What is the structure of this Cap? What are the products of the capping reaction? What enzyme catalyzes the capping reaction?
What is a Cap 0? a Cap 1? a Cap 2? What does "internal methylation" mean? How often does it occur in eukaryotic mRNA?
What is an intron? What is an exon? How are these related to DNA sequences expressed as amino acids in protein species? What does IVS mean?
What is the process of removal of introns from pre-mRNA called? Where does it occur? Where do other eukayrotic mRNA modifications take place? Where does eukaryotic protein biosynthesis occur? How does this differ in mitochondria?
Prokaryotic genes and proteins are collinear. What does this mean? Is this true in eukaryotes? What effect does presence of introns in pre-mRNA have on this statement?
Name three ways that the same DNA sequence can encode different proteins.
Can introns encode proteins? Give an example (see Lewin, Fig. 32.9). How are these proteins useful in recombinant DNA work?
What is "editing" of eukaryotic mRNA? (see Lewin Fig. 32.11)
What is alternative splicing? Give examples of different types of alternative splicing (see Lewin Figs. 6.21, 31.14, 31.16)
What is hetergeneous nuclear RNA? How is it related to eukaryotic mRNA?
What is hnRNP? How is it related to hetergeneous nuclear RNA?
What are the 5 general mechanisms used in RNA splicing?
What are the general principles underlying all splicing mechanisms?
What is a transesterification reaction? What is the attacking group? What common phosphodiester linkage is found in nearly all splicing events? Why is no energy required in a transesterification reaction?
What are the three reactions common to splicing events?
What is the feature common to Group A splicing events?
What is a ribozyme?
What is the dominant structure feature found in Group I introns? Where is nature are Group I introns found?
In the self-splicing reaction for 400 base intron in the 26S rRNA found in Tetrahymena, what are the three steps in the overall reaction?
What are molecule is required, and what does it provide in the first reaction?
At which reaction are the two exons joined together?
What reactions does the excised intron undergo?
What is the G414 and what functions does it provide?
What is the L-19 RNA and how is it related to the intron RNA?
What are the roles of each of the 9 stems in the 9 stem-loop structure of the L-19 RNA?
What is the IGS, and what is its function?
In what sense is the L-19 RNA an enzyme?
What is the relation of the 9 stems to the Substrate Binding Site? to the G-Binding Site? to the catalytic Active Site?
How does this RNA act as an RNA polymerase? what are the reactions? how does it differ from a protein RNA polymerase? how is it similar?
What other catalytic activities do this RNA have?
Why might the endonuclease activity make this RNA a useful RNA restriction enzyme?
How do the kinetic parameters of this RNA as an enzyme compare with those of protein enzymes?
What is RNaseP? What reaction does it catalyze? Is it a ribozyme?
What experiments show that both the RNA and the protein moities of RNaseP are important to its normal function?
How does RNaseP function in pre-tRNA maturation? What other reactions are important in pre-tRNA maturation?
How is the 3'-terminal CCA added to tRNA? What is the function of the CCA?
What is a viroid? what is a virusoid?
What type of splicing reaction is catalyzed by a virusoid? what enzyme catalyzes this reaction?
What is the characteristic RNA secondary structure required for this reaction? What is its name? How is it important in biotechnology? What is the difference between an intramolecular and an intermolecular structure?
In yeast tRNA splicing, what is the importance of secondary structure? what are the important secondary structure characteristics? what is the enzyme responsible for the splicing reaction? what are reactions must also occur in this splicing? what intermediate phosphodiester structure is generated? what happens to this intermediate?
For spliceosome splicing, what is the GU-AG rule?
In splicing, what is the donor site? what is the acceptor site?
What is the major difference in spliceosome splicing from other splicing mechanisms?
In what eukaryotic genes does one find spliceosome splicing?
What are splicing factor proteins? what are small nuclear RNAs? what are snurps? what are snRNPs?
What snurps are used in spliceosome splicing? how do they interact with pre-mRNA?
What is the basic structure of snRNP U1? how does it interact with pre-mRNA? what part of pre-mRNA does it interact with?
What are the 3 stages in a spliceosome splicing reaction?
What is the Branch site? what type of phosphodiester bond is formed at the Branch site? what is the resulting intron structure? what are the critical primary structure features of the Branch site?
What is (are) the role(s) of snRNAP U2 in the splicing reaction? of snRNP U1? of snRNP U5? of snRNP U4 and U6?
What in the structure of U4 and U6 leads to their forming a complex? at what stage in the splicing reaction is this complex disassociated? What takes the place of U4 in a similar complex?
What is a Spliceosome?
How is ATP important in these splicing reactions? Why is ATP not required in other splicing reactions?
In what organisms are Group II introns found? In what type of DNA and genes?
In what ways are splicing of Group II introns similar to that involving spliceosomes? similar to that involving ribozymes?
What features of the Group II intron structure are important for Group II intron splicing?
In the Crick et al experiment:
what was the gene and organism used? Provide reasons why these were good choices.
what was the main drawback or problem with this choice of gene system?
what were the assays used for mutant and wildtype phage?
what was the mutagen used? how does this mutagen work?
+ and - mutants were obtained. What does + and - mean?
with analysis of protein sequences, what does + and - turn out to mean?
what are the phenotypes of ++, --, +1, and -+ double mutants?
what are the phenotypes of triple mutants?
what is suppression? what is intragenic supression? intergenic supression?
what main features of the genetic code were deduced from this experiment?
what main features of the genetic code could NOT be deduced from this experiment?
what experimental results indicated that the genetic code is a triplet code?
In codes:
what is an overlapping code? give an example using the DNA alphabet.
what simple genetic experiment (NOT Crick et al!) shows that the genetic code is not an overlapping code?
what is a comma code? give an example using the DNA alphabet.
what type of "code" is the English language?
The Genetic Code is a triplet code. What comparable statement can be made about the English language?
The statement is made "The genetic code is a universal code." Is this true or false? Why?
The statement is made "The genetic code is a degenerate code." What does this mean?
The statement is made "The genetic code is an unambiguous code." What does this mean?
Is the English language a "degenerate code"? is it an "unambiguous code"?
In making codon identifications, what was the experimental system used by Nirenberg and Matthei?
What was the major problem with this experimental system?
How did the triplet binding assay of Leder and Nierenberg solve this major problem?
What was the triplet binding assay of Leder and Nierenberg?
What are stop codons?
What are sense codons? missense codons? nonsense codons?
How many codons are there? how many of these are stop codons? sense codons? nonsense codons?
How many amino acids are encoded by each codon? What then is this type of code?
How many codons encode each amino acid? What then is this type of code?
What is the translation process?
What are the major reactants in the translation process?
What is the Adaptor Hypothesis of Crick? Why was such a hypothesis needed?
What is the Adaptor Molecule?
What are some unique features of the primary structure of tRNA molecules?
What is Pseudo-U ( Y )? how is it formed? what is the structure of DiHydro-U?
Roughly, how many different types of tRNA molecules are there?
What are the major features of tRNA secondary structure?
What are the functions of each of the "arms" of a tRNA molecule?
What is the 3-D structure of tRNA molecules?
What types of unusual base pairing occurs in the tRNA 3-D structure?
What are the two main functions of tRNA molecules?
How does the tRNA structure physically separate these two functions?
What are the differences between Type1 and Type 2 tRNA molecules?
What is the difference between "charged" tRNA and "uncharged" tRNA molecules?
What is an AminoAcyl-tRNA synthetase?
What are the two classes of such synthetases? How do these each interact with tRNA molecules? How is the tRNA structure distorted in these interactions?
What are primary structure differences in the two classes of synthetases?
What are the two steps in the amino-acylation reaction? What is the role of ATP in these reactions?
What does "proofreading" mean?
What are the proofreading mechanisms used by aminoacyl-tRNA synthetases in the above 2 reactions?
Which of these mechanisms are at the tRNA level and which are at the amino acid level?
Why is proofreading less important in protein biosynthesis than in DNA replication?
What is the "anticodon"? How does it interact with the "codon"?
What is the Wobble hypothesis? Why was it put forward, ie what does it explain?
What is the key new base pairing permitted in the Wobble hypothesis?
Was it Wobble who proposed this hypothesis? If not, who?
How effective is the Wobble hypothesis in considerations of genetic code degeneracy?
How is degeneracy of the code useful to organisms of variable %(A+T)?
What is the variation in %(A+T) found in organisms throughout the world?
What is the variation in %(A+G) found in organisms throughout the world?
What is a suppressor tRNA?
What is a nonsense mutation? How "leaky" are these mutations?
What are amber mutations? ochre mutations? opal mutations?
How does suppression by a suppressor tRNA molecule work?
Why doesn't this suppression work when it shouldn't, ie on nonmutant codons?
Understand the difference between nonsense and missense suppression.
What is a missense mutation? How "leaky" are these mutations?
What does tRNA suppression have to do with ambiguity of the code?
What is frameshift suppression?
How can frameshift suppression be used to control level of expression of genes? Give 2 examples.
What are tRNA gene clusters? How are these relevant to tRNA gene expression?
What are ribosomes?
What are the 6 ribosome species found in organisms?
What are rRNA species? Which of these species are found in each of the ribosome subunits?
Roughly how many proteins are found in each of the ribosome subunits?
How well is the detailed structure of the ribosome known?
How well is the detailed structure of 5S rRNA known?
How well is the detailed structure of 16S rRNA known?
How are ribosomes made, ie what are features of the assembly process?
What are split proteins?
What is the major function of rRNA molecules?
What other functions are known for some of the rRNA molecules?
Do any rRNA molecules participate in enzymatic reactions associated with protein biosynthesis?
Do any tRNA molecules participate in enzymatic reactions associated with protein biosynthesis?
What are the active sites found on each of the ribosomal subunits?
What are the two major functional parts of a tRNA molecule?
How does the structure of a tRNA molecule effectively separate these two functions in space?
Where are the active sites on the ribosomal subunits for these two functions?
What do these two functions have to do with the Adaptor hypothesis?
What is the Exit site on the large ribosomal subunit?
What is your reaction to the placement of this site in Lewin, Fig. 9.11, relative to the P and A sites?
What is the P site?
What is the A site?
What does peptidyl transferase do?
What is the molecule which has peptidyl transferase activity?
In general, how has genetics been useful in delineation of roles of ribosomal proteins, specific parts of rRNA molecules, and of protein biosynthesis protein cofactors in specific steps in protein biosynthesis, either complex formation and molecular interactions or in catalytic activities?
In general, how have specific drugs and antibiotics been useful in similar delineations?
How does translation fit into the general scheme called the Central Dogma of Molecular Biology?
What are the three major steps in the translation process?
What are the four major molecular constituents for protein biosynthesis?
Which ribosomal subunits exist in pools in prokaryotic cells?
Which of these ribosomal subunits are active in protein biosynthesis?
What is the direction of assembly of proteins?
What is the direction of reading of the mRNA?
What is the direction of transcription?
What is the rate at which proteins are assembled, ie the rate at which ribosomes read mRNA?
What is the rate of prokaryotic transcription?
What is the rate of DNA replication?
How does all this information fit with a coordinated transcription-translation process in prokaryotes?
What is meant by a coordinated transcription-translation process in prokaryotes?
What are polyribosomes? How can they be isolated? What is the evidence for their existence?
Design a CsCl density labeling experiment that would show that the 70S ribosome "turns over", ie is dissociated into its 30S and 50S subunits, between each protein biosynthesis event.
How many ribosomes are there per bacterial cell?
How do you think this number varies with growth rate of the bacterial cell?
What fraction of the bacterial cell mass is accounted for by ribosomes?
What are the three IF factors required for the initiation process of protein biosynthesis in prokaryotes?
What is an IF factor?
What are the four major constituents that come together to form the final translation initiation complex?
Which interact first in prokaryotes, fMet-tRNAf and 30S ribosomes, or mRNA and 30S ribosomes?
What is the Shine-Delgarno sequence? Where is it found? How is it related to the start codon for translation? What is the start codon for translation?
How is the Shine-Delgarno sequence used? What complex is formed as a result of this "usage"?
How is the Shine-Delgarno sequence related to a "ribosome-binding site" on the mRNA?
What tRNA molecule is used in prokaryotic translation initiation?
How does this tRNA molecule differ in structure from other tRNA molecules?
What unique characteristics as a tRNA molecule doe these structure properties confer on this "initiation" tRNA molecule?
What is "fMet"? What enzymatic reaction creates "fMet"? What is the "f" in "fMet"?
Know how to draw the structure of fMet.
What is the function of IF-3? of IF-1?
What does IF-2 do? What cofactor is required in IF-2 activity? What is the role of this cofactor? What is the fate of this cofactor, ie what happens to it?
What is the final step in the initiation process for prokarytic translation?
What are the constituents of the final initiation complex, and how are they bound to each other?
What happens to the "f" of "fMet" during growth of the nascent polypeptide chain?
What is meant by "nascent polypeptide chain"?
What is the cognate molecule for fMet-tRNAf in eukaryotic translation initiation?
What are the eukaryotic cognate eIF factors to the three prokaryotic IF factors?
Which interact first in eukaryotes, Met-tRNAi and 40S ribosomes, or mRNA and 40S ribosomes?
What eIF factors are involved in the interaction of mRNA and 40S ribosomes?
What is the cognate sequence to the prokaryotic Shine-Delgarno sequence?
How does the eukaryotic initiation complex find the mRNA start codon? What cofactor is involved in this process? What is the role of this cofactor and what is its fate?
How does the eukaryotic start codon(s) compare with the prokaryotic start codon(s)?
What eukaryotic initiation complex finds the mRNA start codon?
Which eIF factor is involved in the Met-tRNAi - 40 S ribosome interaction with eukaryotic mRNA? What cofactor is also required for this interaction? What is the fate of this cofactor?
What events occur on the eukaryotic mRNA prior to binding of the Met-tRNAi - 40 S ribosome complex? What eIF factors are involved in these events? What is CAP binding protein? What role does ATP play in these events?
What are the steps in formation of the final eukaryotic initiation complex? What eIF factors are involved and what do they do? What are the constituents of this final complex and how do they interact with each other?
What are EF factors?
What are the three prokaryotic EF factors? Which are functional in the translation elongation process?
What is the function of EF-Tu? What cofactor does EF-Tu use, and what is its fate?
What is the active form of EF-Tu and what is the inactive form?
What happens to EF-Tu when it is released from the 70S ribosome?
What is EF-Ts and what is its function?
What are the relative numbers of EF-Tu and EF-Ts per bacterial cell? How do these numbers compare with the number of ribosomes per cell?
What is (are) the cognate eEF factors in eukaryotes to the prokaryotic EF-Tu and EF-Ts factors?
Eukaryotic factor eEF-1 is often called EF-T. Why might this be the case?
What are the roles of charged tRNA molecules in the P and A sites of the 70S ribosome in peptide bond formation?
What is the reaction that occurs in peptide bond formation? What enzymatic activity catalyzes this reaction?
What is puromycin and how does it work?
What is the translocation step in translation elongation?
What is the energy source for translocation?
What molecules are translocated relative to which other molecules?
What are the two steps in translocation?
What EF factor(s) is (are) required for this translocation in prokaryotes? in eukaryotes?
How is binding of EF factors involved in coordinating the steps of the elongation process?
What, in summary, are the steps in the elongation process? Why is this process considered to be a cyclic process? Why is it considered to be a sequential process?
Why is the prokaryotic EF-G factor sometimes called a "GTP-dependent translocase"?
What is the signal for cessation of the elongation process and start of the termination process?
How can "premature termination" occur? How can termination NOT occur at the correct signal?
What are the prokaryotic RF factors?
What does RF mean?
What signals do each of the RF factors recognize?
What events occur as a result of RF activity?
What two differences are there between prokaryotic termination and eukaryotic termination?
What is an ORF? What is an URF? Why are there 3 reading frames on mRNA? Why are there 6 reading frames in a DNA sequence?
Are more than one reading frame ever used in protein biosynthesis? This case is sometimes called "overlapping genes". Why is this the case?
Does transcription off BOTH DNA strands in the same DNA sequence ever occur?
Which eukaryotic RNA polymerase transcribes most eukaryotic genes encoding proteins?
What are the two types of DNA sites recognized by the RNApolII Basal Transcription Apparatus?
What are Upstream Elements? How do they differ from the above two types of DNA sites? Where are they found? What is the structure of the Upstream Elements?
What types of molecules recognize Upstream Elements? What is(are) the function(s) of these molecules? How do they execute these function(s)?
What are some examples of Upstream Elements? of the molecules that recognize Upstream Elements?
What is the effect on transcription of mutations within the Upstream Elements?
Do these Upstream Elements and their recognition molecules show specificity for particular types of tissue? types of organisms? types of genes?
How important is the precise position and orientation of Upstream Elements? What do these properties say about the mechanism of interaction between the recognition molecules and RNApolII?
Can Upstream Elements be recognized by more than one molecule? Are these molecules always/never Activators? always/never Repressors? If some are Repressors, how do they differ from prokaryotic Repressors? how are they similar to prokaryotic Repressors?
What are Enhancers? Where are they found? Is their activity orientation dependent? To be active, must enahncers be on the same DNA molecule as the transcribed gene? What do these properties indicate regarding mechanism of their activity? What are Enhancers called in yeast?
What is the effect on transcription of mutations in Enhancers? Does such effect depend on where within the Enhancer the mutation is located? What does this say about the structure within the Enhancer? How do the structural elements or motifs found within Enhancers compare with those found within promoters? Understand Lodish, Figure 11-41.
What is a Reporter Gene? How is a reporter gene used to assay mutations in transcription factor binding sites? How is such information used to define such binding sites? How can one use such binding sites to identify cognate transcription factor proteins? How can one use this methodology to purify the cognate transcription factor protein? How can the protein be used in turn to better define the binding site(s)? What is the two-plasmid assay system for transcription factor activity? Understand Lodish, Figs 11-43, 11-44, 11-45.
What is the function of Enhancers? What types of molecules recognize Enhancers? How do these molecules interact with Enahncers? What structural motifs do these molecules have and how do these motifs function in interactions with the Enhancers? Understand the major features shown in Lodish, Figs. 11-48, 11-50, 11-51.
How do these molecules interact with RNApolII? What are the functions of these interactions?
What is a multiple sequence alignment? What is the value of the multiple sequence alignment shown in Lodish, Fig. 11-49?
What is a Heterodimeric Transcription Factor? How do such increase regulational diversity? Understand Lodish, Fig. 11-52.
What are Response Elements? How are they similar to, and how do they differ from, Upstream Elements? Enhancers? What do these Response Elements respond to? What are nucleotide direct repeats? inverted repeats? How are such repeats relevant to Response Elements? (see Lodish, Fig. 11-59).
What is the Heat Shock Transcription Factor? What does it do? How does it do this? How is it similar in function to the prokaryotic heat shock sigma factor? how does it function differently from this sigma factor?
What is the metallothionein (MT) gene? How is it activated? Which response elements are found in its regulatory region? Are all of these found in the promoter? Do the response elements function independently? What do they respond to?
What is the GRE? What molecule binds to the GRE? What molecules do the GRE respond to?
What are the two functions of Transcription Factors? How are the parts of the protein that execute each of these functions separated in the structure of the protein? What type of experiments shows this separation?
What are the functions of the GAL4 transcription factor? What DNA site does GAL4 bind to? What proteins does GAL4 bind to? What is the structure of the GAL4 protein vis a vis its functions?
What is the GAL80 protein and what is its function? What regulatory feature or concept does GAL80 exemplify?
What is the GAL4-LexA chimeric transcription factor experiment, and what does this experiment show?
What are the most common DNA binding domain motifs found in Transcription factors? How do these motifs interact with DNA? What are some of the transcription activation motifs found?
Compare the Enhancer-Transcription Factor mechanism for gene regulation found in eukaryotes with the Repressor-Operon mechanism for gene regulation found in prokaryotes. What are similarities? What are differences?
How to the features of regulation of expression of the mouse transthyretin (TTR) gene (Lodish, Fig. 11-56) and of gene expression regulation by the protein encoded by the Wilms' tumour (WT1) gene (Lodish, Fig. 11-55) exemplify the above concepts?
What is a lipid-soluble hormone? Give some examples. How do such hormones enter a eukaryotic cell? Would they be able to enter a prokaryotic cell by the same mechanism?
What new protein domain is found in Transcription Factors that respond to small molecules such as lipid-soluble hormones (see Lodish, Fig. 11-60)? What then are the three functional domains found in such transcription factors?
What is a fusion protein? How was such used to demonstrate effects of hormone binding by the glucocorticoid receptor protein? (see Lodish, Fig. 11-61). How does this type of experiment DIFFER from the two-plasmid reporter gene experiment (see Lodish, Fig. 11-45)? How are the two types of experiments SIMILAR?
In the model for hormone-dependent gene activation by the glucocorticoid receptor, why is the receptor thought to be bound to an inhibitor in the absence of hormone? (see Lodish, Fig. 11-62)
What is Signal Transduction? With what types of hormones and small molecules is signal transduction needed? Why? Understand Lodish, Fig. 20-2; compare Lodish, Fig. 11-57 and Table 20-2.
How is Signal Transduction related to cell-cell communication?
What are the two most common mechanisms whereby signals are "transduced" to proteins within a cell? What are examples of "second messengers"? What are "second messengers"? See Lodish, Fig. 20-3.
What are the four classes of cell-surface receptors for signal transduction? How do they differ from each other? Understand the basic features of Lodish, Fig. 20-4.
What is affinity labeling? How is affinity labeling used to assay a receptor protein (see Lodish, Fig. 20-8)? How does affinity labeling differ from affinity chromatography? How can affinity labeling be used to purify a cDNA clone from a cDNA library harboring the receptor gene (see Lodish, Fig. 20-9)? How would this cDNA clone be used?
What is the JAK-STAT signal transduction pathway? What is JAK, and what is its function? What is STAT and what is its function? Are there any second messengers in this pathway? How does this pathway differ from that of a lipophillic hormone such as a glucocorticoid steroid?
How do the data shown in Lodish, Fig. 11-63, show that phosphorylation of a specific tyrosine residue in Stat91 is essential for INF-gamma signal transduction? Why were cells that do not normally express Stat91 used? What was the assay that was used?
Is the statement "The JAK-STAT model shown in Lodish, Fig. 11-64, is essentially a simplified version of that shown in Lodish, Fig. 20-52" true? What are the common features of the two models? What are the main differences between the two models? How is cell specificity provided in cellular response to different interferons? What is an SH2 domain and what is its function?
What is the basic "two membrane proteins - G protein" mechanism of G-Protein-linked Receptors? see Lodish, Fig. 20-4a. What is an "effector enzyme"?
What is an agonist? What is an antagonist? What are these compounds used for?
What is the basic structure of a G protein - linked Receptor? Why are these often called 7TM proteins? How is isoproterenol bound to the receptor (see Lodish, Fig. 20-13)?
What is the structure of cyclic-AMP? Why is cyclic-AMP called a "second messenger"? How is it made as a second messenger?
What is the function of adenylate cyclase? What is its structure (Lodish, Fig. 20-15)?
What is the subunit structure of a G-protein? What happens to the G-protein when the receptor is activated? Why is it called a "G-protein"? What happens to each subunit of the G-protein during activation of adenylate cyclase? See Lodish, Fig. 20-16.
How does Ras protein cycle between an active GTP-bound form and an inactive GDP-bound form (see Lodish, Fig. 20-17)? What is the GTPase protein superfamily? How does this Ras protein cycle serve as a model for similar G-protein cycling? What is the GEF protein? What is the GAP protein? What are the Ras-like and GAP-like domains of GS alpha and how do they function in GDP-GTP exchange (Lodish, Fig. 20-18)?
What is the difference between GS and GI proteins (see Lodish, Fig. 20-20)?
How does cAMP function in regulation of cellular metabolism? What is a cAMP-dependent protein kinase (PKA)?
How does cAMP concentration influence glycogen metabolism (see Lodish, Figs. 20-25, 20-26)?
What is the "kinase cascade"? How does the kinase cascade amplify the hormone signal (see Lodish, Fig. 20-27)?
What are CRE and CREB? How do they function in the cAMP regulatory response? see Lodish, Fig. 20-51a.
What two major mechanisms are used to down regulate hormone-activated receptors? What is a BARK and how does it function (see Lodish, Fig. 20-50)?
What is the basic receptor tyrosine kinase (RTK) signal transduction mechanism (see Lodish, Figs. 20-4c, Fig. 20-38)?
How does an active receptor tyrosine kinase form upon binding receptor ligand (see Lodish, Fig. 20-28)?
What are the structural characteristics of a RTK molecule?
What are examples of ligands that activate an RTK?
What reactions are catalyzed by active RTK molecules?
What is an SH2 domain and how does it function (see Lodish, Fig. 20-30)? How are SH2 domains important vis a vis interactions with active RTK molecules? How does an SH2 domain distinguish between an active and an inactive RTK?
How is the signal that activates an RTK transmitted to Ras molecules (see Lodish, Fig. 20-32)? What is an active Ras molecule?
How does GRB2 function in RTK signal transduction? What is an SH3 domain (see Lodish, Fig. 20-35) and how does it differ from an SH2 domain? How is GRB2 an "adaptor molecule"? How are such "adaptor molecules" important in apoptosis (see Death Receptor journal article and DD vs DED domains)? What is the Sos protein and how does it function in RTK signal transduction?
What is the "protein kinase cascade" that transmits signals from activated Ras downstream to the nucleus (see Lodish, Fig. 20-36)? What does activated MAP kinase do?
How was the "two-hybrid system" used to show that Ras protein interacts directly with Raf protein? What is the "two-hybrid system" (see Lodish, Fig. 20-37)? What is the "bait" plasmid? What is the "fish" plasmid? Why are yeast cells used? Why are these cells auxotrophic for THREE biosynthetic markers, e.g. histidine, leucine, and tryptophane?
How can activated MAP kinase lead directly to transcription
activation of response genes (see Lodish, Fig. 20-51b)
The complete genome sequences are now known for what types of prokaryotes? What is the genome of these organisms? Genome sequencing is in progress for what other prokaryotes?
Browse through the Web pages set up for some of these organisms at the TIGR Microbial Database site.
What is TIGR?
Other prokaryotic genomes have been sequenced, but the data are proprietary to certain companies. How are these data being used for commercial profit? What prokaryotic organisms might you guess are included here?
What eukaryotes are currently the "model organisms" for which whole genome sequencing programs are in progress? Have any of these been completed? which?
Given a new DNA sequence, what is the first step in computerized study of the sequence?
Why is there such a high percentage of database matches for the Mycoplasma genitalium genome (92%), and such a comparatively low percentage of database matches for the Methanococcus jannaschii genome (44%)? What is an ECE? What is a "database match"?
What is "genomics"? What do you think the following are: "functional genomics"? "structural genomics"?
What are Lander's "Ten Goals" for human genomics?
What are sequence databases? organism databases? "smart"
databases?
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