cellular Flashcards
regulators of cell cycle
cyclins, CDKs, tumor suppressors
shortest phase of cell cycle
MITOSIS: prophase. metaphase. anaphase. telophase.
cyclins
regulatory proteins.
phase-specific.
ACTIVATE CDKs.
CDKs (cyclin-dependent kinases)
constitutive and inactive.
tumor suppressors
Rb and p53.
inhibit G1 to S progression.
*mutations = unrestrained growth
permanent cells
remain in G0.
regenerate from STEM CELLS.
neurons.
skeletal m.
cardiac.
RBCs.
stable (quiescent) cells
enter G1 from G0 when stimulated.
hepatocytes.
lymphocytes.
labile cells
never go to G0.
rapidly divide with short G1.
BM.
gut epith.
skin.
hair follicles.
RER
site of synth of secretory (EXPORTED) proteins.
N-linked oligosaccharide addition to many proteins.
which cells are rich in RER?
mucus-secreting GOBLET CELLS of small intestine.
Ab-secreting PLASMA CELLS.
Nissl bodies
RER of neurons-
synthesize enzs and peptide NTs.
free ribosomes
unattached to any memb.
synth of cytosolic and organellar prots.
SER
- steroid synth (hormones).
- detox of drugs, poisons.
- carb metabolism.
which cells are rich in SER?
liver hepatocytes.
steroid-producing cells of adrenal cortex.
peroxisome
catabolism of VLCFA (very long chain fatty acids) and amino acids
proteasome
barrel-shaped protein complex.
degrades damaged/unnecessary proteins tagged with UBIQUITIN for destruction.
Golgi apparatus
distribution center for proteins and lipids-
from ER to plasma memb and vesicles.
how does Golgi change asparagine?
modifies N-oligosaccharides
how does Golgi change serine and threonine?
adds O-oligosaccharides
mannose 6-phosphate
Golgi adds mannose 6-phosphate to proteins for trafficking to LYSOSOMES
endosome
sorting center for material from outside cell or from Golgi - sends it to lysosomes for destruction or back to memb/Golgi for further use
vesicular trafficking prot: COP I
retrograde.
Golgi to ER.
vesicular trafficking prot: COP II
anterograde.
RER to cis-Golgi.
vesicular trafficking prot: clathrin
trans-Golgi
to
lysosomes, memb
to endosomes (receptor-mediated endocytosis)
I cell disease
(inclusion cell disease)
inherited lysosomal storage disorder.
fail to add mannose 6-phosphate tag to lysosome proteins = enzs secreted outside of cell instead of being targeted to lysosomes.
features of I cell disease
coarse facial features.
CLOUDED corneas.
restricted joint mvmt.
high plasma levels of lysosomal enzymes.
*often fatal in childhood
microtubule
cylindrical structure.
helical array of polymerized dimers (ALPHA and BETA TUBULIN) - each dimer has 2 GTP.
microtubules are incorporated into…?
flagella (dynein).
cilia (dynein).
mitotic spindles.
axoplasmic transport in neurons*
molecular motor proteins
transport cellular cargo (organelles, intracellular vesicles) toward opposite ends of microtubule tracks.
use ATP.
dynein
Retrograde to microtubule (+ to -)
kinesin
Anterograde to microtubule (- to +)
drugs that act on microtubules
Mebendazole, thiabendazole. Colchicine. Griseofulvin. Vincristine, vinblastine. Paclitaxel.
“Microtubules Can Grow Very Progressively”
Chediak-Higashi syndrome
microtubule polymerization defect =
decreased fusion of phagosomes and lysosomes.
results in recurrent pyogenic infxs, partial albinism, peripheral neuropathy.
cilia
9+2 arrangement of microtubules.
contains dynein.
dynein in cilia
axonemal dynein.
ATPase links peripheral 9 doublets and causes bending of cilium by differential sliding of doublets.
Kartagener’s syndrome
IMMOTILE CILIA due to dynein arm defect.
results in male and female infertility (sperm immotile), bronchiectasis, recurrent sinusitis.
assoc. with situs inversus.
cytoskeleton: actin, myosin
microvilli.
muscle contraction.
cytokinesis.
adherens junctions (intermediate).
cytoskeleton: microtubules
movement:
cilia. flagella. mitotic spindle. axonal trafficking. centrioles.
cytoskeleton: intermediate filaments
structure:
vimentin. desmin. cytokeratin. lamins. GFAP. neurofilaments.
plasma membrane
asymm lipid bilayer: cholesterol 50%. phospholipids 50%. sphingolipids. glycolipids. proteins.
high chol or long saturated FA content of plasma memb means….?
increased melting temp.
decreased fluidity.
vimentin
conn tissue
desmin
muscle
cytokeratin
epith cells
GFAP
neuroGlial cells (astrocytes)
neurofilaments
neurons
Na-K ATPase
ATP site on cytoplasmic membrane:
1 ATP consumed.
3 Na out.
2 K in.
ouabain
inhibits Na-K ATPase by binding to K site
cardiac glycosides (digoxin, digitoxin)
directly inhibit Na-K ATPase
= indirect inhibition of Na/Ca exchange
(increase intracellular Ca = increase cardiac contractility)
most abundant protein in human body?
collagen
collagen function
organizes and strengthens ECM
type I collagen
90%. bone skin tendon dentin fascia cornea late wound repair (scars)
type I collagen defect in…
osteogenesis imperfecta
type II collagen
cartilage (including hyaline)
vitreous body
nucleus pulposus
type III collagen
AKA reticulin. skin blood vessels lungs intestine RES uterus fetal tissue granulation tissue
type III collagen defect in…
Ehlers-Danlos
type IV collagen
basement memb
or basal lamina
type IV collagen defect in…
Alport syndrome
collagen formation step 1: synthesis
in RER:
translate collagen alpha chains (preprocollagen).
usually GLY-X-Y polypeptide where
X and Y are proline and lysine.
collagen formation step 2: hydroxylation
in RER:
hydroxylate specific proline and lysine residues.
REQUIRES VIT C
collagen formation step 3: glycosylation
in RER:
glycosylate pro-alpha-chain hydroxylysine residues.
form PROCOLLAGEN via hydrogen and disulfide bonds (TRIPLE HELIX of 3 pro-alpha-chains)
collagen formation step 4: exocytosis
procollagen triple helix exocytosed into extracellular space
collagen formation step 5: proteolytic processing
outside fibroblast:
cleave terminal regions of procollagen to transform into insoluble TROPOCOLLAGEN
collagen formation step 6: cross-linking
outside fibroblast:
reinforce numerous tropocollagen molecules with covalent lysine-hydroxylysine cross-linkage by LYSYL OXIDASE to make COLLAGEN FIBRILS
what step in collagen formation does scurvy affect?
vit C deficiency — affects hydroxylation of proline and lysine residues
osteogenesis imperfecta
genetic bone disorder (BRITTLE BONE) caused by various gene defects.
most commonly auto dom with abnormal collagen type I.
features of osteogenesis imperfecta
- multiple fractures with minimal trauma.
- BLUE SCLERA due to translucency of conn tissue over choroid.
- hearing loss due to abn middle ear bones.
- dental imperfections due to lack of dentin.
may be confused with child abuse.
which form of OI is most fatal?
type II - in utero or neonatal period
Ehler-Danlos syndrome
faulty collagen synthesis.
varying inheritance and severity.
auto dom or recessive.
what type of collagen is most often affected in Ehler-Danlos syndrome?
type V (classic syndrome)
*6 types total
features of Ehler-Danlos syndrome
- hyperextensible skin.
- tendency to bleed (easy bruising).
- hypermobile joints.
may be assoc with joint dislocation, berry aneurysms, organ rupture.
Alport syndrome
due to various gene defects leading to abnormal TYPE IV collagen.
most common is X-linked recessive.
features of Alport
- progressive hereditary nephritis
- deafness
- ocular disturbances
*type IV important struct component of BM in kidney, ears, eyes
elastin
stretchy protein in lungs, large arteries, elastic ligaments, vocal cords, ligamenta flava.
ligamenta flava
connect vertebrae: relaxed and stretched conformations
elastin is rich in which AA?
proline. glycine. lysine. alanine. valine.
(nonpolar, non-hydroxylated forms)
tropoelastin
elastin precursor.
has fibrillin scaffolding.
what breaks down elastin?
elastase
what inhibits elastase?
alpha 1 antitrypsin
wrinkles of aging
due to reduced collagen and elastin prod.
Marfan’s syndrome
defect in fibrillin
emphysema
can be caused by alpha 1 antitrypsin deficiency resulting in EXCESS ELASTASE activity
PCR purpose
amplify desired fragment of DNA
PCR steps
- denaturation.
- annealing.
- elongation.
*repeated multiple times for DNA sequence amplification (25-35 cycles)
PCR: denaturation
DNA denatured by heating to generate 2 separate strands
PCR: annealing
during COOLING, excess premade DNA primers anneal to specific seq on each strand to be amplified
PCR: elongation
heat-stable DNA polymerase replicates the DNA seq that follows each primer
components needed for PCR
- DNA template (seq may be unknown).
- two primers complementary to regions flanking DNA template.
- heat-stable DNA polymerase that replicates target sequence between primers.
- deoxynucleiotide triphosphates to build new DNA.
agarose gel electrophoresis after PCR
for SIZE separation of PCR products (small molecules travel further).
compared against DNA ladder.
Southern blot
DNA
Northern blot
RNA
Western blot
protein
Southwestern blot
DNA-binding proteins (TFs, nucleases, histones)
microarray
thousands of nucleic acid sequences arranged in grids on glass or silicon.
DNA/RNA probes are hybridized to chip.
scanner detects relative amts of COMPLEMENTARY BINDING.
purpose of microarray
to profile gene expression levels of thousands of genes simultaneously.
can detect SNPs.
ELISA
enzyme-linked immunosorbent assay.
rapid immunologic test for AG-AB REACTIVITY.
ELISA with test Ag
Ag coupled to color-generating enzyme.
see if imm system recognizes Ag.
(if target is acquired in sample, test soln will have intense color rxn to indicate positive result)
ELISA with test Ab
Ab coupled to color-generating enzyme.
see if a certain Ag is present.
(if target is acquired in sample, test soln will have intense color rxn to indicate positive result)
Sn and Sp of ELISA
both approach 100% though FP and FN possible
FISH
fluorescence in situ hybridization.
fluorescent DNA or RNA probe binds specific gene site of interest on chromosomes.
FISH use
for specific localization of genes.
direct visualization of anomalies.
no fluorescence in FISH
= gene has been deleted
what phase are chromosomes used in karyotyping in?
METAPHASE
karyotype can be performed with samples of…?
blood
BM
amn fluid
placental tissue
karyotype use
diagnose chromosomal imbalances
cloning
prod of recombinant DNA molec that is self-perpetuating
what is the first component of cloning?
euk mRNA of interest
what enzyme acts on mRNA in cloning?
reverse transcriptase produces cDNA from mRNA
what is used to produce a cDNA library?
insert cDNA into bacterial plasmids containing antibiotic resistance genes - those that survive on Abx medium produce library
transgenic strategies
to modify gene expression (in mice) by:
- random insertion of gene into mouse genome (constitutive).
- targeted insertion or deletion of gene through homologous recombination with mouse gene (conditional).
Cre-lox system
can inducibly manipulate genes at specific development points using Abx-controlled promoter
RNAi
dsRNA complementary to mRNA sequence of interest - transfected into humans and promotes degradation of target mRNA (thus knocking down gene expression)
