Lecture 41: Protein Synthesis Control/Cancer

Question 1

Mitochondrial protein translocation process

Answer 1

1. OMM receptor binds pre-seq. helix
2. TOM complex forms; OMM/IMM channels align
3. Protein threads through to matrix
4. Mitochondrial matrix chaperones mediate folding, translocation

Question 2

Mitochondrial localization sequence

Answer 2

Pre-sequence, amphiphilic helix w/ one hydrophobic side one charged side

Question 3

PDH defect clinical case

Answer 3

High lactate due to PDH E1alpha not being imported efficiently into mitochondria; Pro substitution mutation disrupts pre-sequence localization helix for receptor

Question 4

Nucleus selective diffusion barrier

Answer 4

Small things pass thru nuclear membrane, larger proteins require active transport. Mediated by Nup (nucleoporin) proteins e.g. Phe-Gly (FG) Nups

Question 5

Nuclear Localization Signal

Answer 5

Lys-Lys-Lys-Arg-Lys
NLS binds cytoplasmic importins for transport into nucleus via nuclear pores

Question 6

Importins/exportins

Answer 6

Carriers that mediate transport into/out of the nucleus by binding w/ Nups in nuclear pores

Question 7

Nuclear lamina

Answer 7

Thick layer of intermediate filaments (nuclear lamins) that associate, not attach to, nuclear envelope. Maintains nuclear structure and stabilizes nuclear pore complexes

Question 8

Hutchison-Gilford Progeria Syndrome

Answer 8

Nuclear lamina defect due to truncated modified progerin attaching nuclear lamina to envelope. Inhibits import/export from nucleus. Treatment w/ farnesyl transferase inhibitors (farnesyl group is what attaches mutant protein to envelope, normally cleaved)

Question 9

Restriction endonucleases (enzymes)

Answer 9

Perform seq-specific cleavage of 4, 6, or 8 bases at double strand palindromic sites. Require perfect base pairing to function

Question 10

Restriction enzyme sites

Answer 10

1. Blunt ends (cleavage in middle)
2. Sticky ends (leaves 3’ or 5’ overhang)
   All are palindromes over 2 strands

Question 11

Restriction Fragment Length Polymorphism (RFLP)

Answer 11

Technique that uses restriction enzyme specificity to distinguish normal from mutant genes. Requires known mutation e.g. w/ mutant, peptide is not cleaved into fragments on southern blot

Question 12

Proto-oncogene

Answer 12

Normal gene with potential to become oncogene

Question 13

Oncogene

Answer 13

Gene for a protein that promotes cancer development when deregulated; typically hyperactivates cell properties and has gain of function mutation e.g. MAP kinase signaling cascade

Question 14

Tumor suppressor gene

Answer 14

Encodes protein that protects from cancer development; usually loss of function in cancer

Question 15

Chromosomal instability pathway to cancer

Answer 15

As mutations accumulate, genomic stability decreases, resulting in further mutations and increasing chance for double hit

Question 16

Microsatellite instability pathway

Answer 16

Methylation of tumor suppressor promoters due to upregulation of genes that methylate.

Question 17

MDM2

Answer 17

Protein that mutually regulates p53; MDM2 downregulates p53 and p53 upregulated MDM2. Under stress, MDM2 dissociates from p53 allowing p53 to exert downstream effects (checkpoint activity). Many cancers upregulate MDM2 accelerating p53 degradation.

Question 18

CRISPR-Cas9

Answer 18

Cas9 recognizes DNA fragments in CRISPR sequences and uses the guide RNA reference to excise the corresponding DNA from the genome. KO can be done by repairing genome w/ NHEJ or homologous template can be provided for knock-in through homologous recombination.