focus cards Flashcards
what are the differences between SEM,DIC, and TEM
SEM: electrons hit off the surface of the fixed, dehydrated, metal-coated cell, they show 3D structures in high quality
DIC: uses polarized light and optical interference to enhance contrast from transparent slides of live/unstained cells to produce 3D-like images
TEM: passes electrons through a thin sample to create images based on electron density, high-res, but 2D images of internal structures
what is the structure of AF?
is a monomer, helix-like format, ATP used to bind together monomers into polymers, has a plus (growth) and minus (decay) end
what is treadmilling in actin filaments?
when individual filaments are getting added and removed from the positive and negative ends of actin equally, giving the illusion of movement in the direction of the plus end
what is the relationship between rate of hydrolisis and critical concentration (Cc)
above critical concentration, addition of subunits is faster than hydrolysis, whereas below, hydrolysis overpowers. at the minus end, subunits are still being added but hydrolysis catches up and allows for decay
what is myosin ii? describe its structure.
myosin ii is a motor protein for cell movement and muscle contraction
coiled coil of 2 alpha helices form a heavy chain, which in turn, form tails and heads + two smaller chains attached to each heavy chain, they stabilize and amplify conformational changes in myosin during movement, stabilie the neck and regulate activity
- head: is a catalytic region responsible for ATP hydrolysis and force generation for movement
- tail: responsible for dimerization (pairing with other myosin) and interaction with cargo/structural elements
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what is the structure of a sarcomere?
- actin (thin) filaments = g-actin (globular) + f-actin (helical), attach to z-disc and are cross-linked by proteins, minus ends aligned towards m-line, plus ends towards zdiscs with cap z on them
- myosin (thick) filaments = binds to actin and generates force using ATP, centered in the middle of the sarcomere, anchored by M-line
- capZ = anchors actin at z-discs, stabilizes them, prevents loss/addition (alpha and beta subunits create it)
- z-disc (zline) = boundary of each individual sarcomere, provides attachment points for actin and connects sarcomeres end-to-end, made of alpha actinin + capZ
- m-line= central point of sarcomere when myosin is anchored, helps maintain myosin arrangement
what are the stages of the cell cycle (in order with subphases)
interphase (g1>s>g2), m phase (prophase>metaphase>anaphase>telophase), cytokinesis
explain what takes place in interphase and all its substeps
g1: cell grows, synthesizes proteins, makes organelles, prepares for DNA replication
s: DNA replicated, doubles material, chromosomes form sister chromatids connected at centromeres
g2: cell grows and preps for mitosis, proteins and organelles needed for division are produced
explain what takes place in the m phase and all its substeps
- prophase: chromosomes condense, nuclear envelope breaks down, mitotic spindle forms
- metaphase: chromosomes align at the metaphase plate, spindle fibers attach to sister chromatids’ centromeres
- anaphase: sister chromatids are pulled apart by spindle fibers to opposite poles of cells
- telophase: chromsomes decondense at poles, nuclear envelope reforms around the sets of chromosomes
explain what occurs during cytokinesis
cytoplasm divides in 2, forms genetically identical daughter cells, cleavage/cell plate forms
what are the checkpoints in the cell cycle?
between G2 and M: DNA replication checkpoint (is all dna replicated and is the environment favourable)
between metaphase and anaphase in M: spindle attachment checkpoint (are all chromosomes attached to the spindle)
end of G1 to beginning of S: DNA damage checkpoint (is environment favourable)w
what does the dna damage checkpoint do
p53 halts the cycle, initiates DNA repair and apoptosis. assesses integrity of dna, if damaged and subsequently repairs said damaged dna, if damage is irreparable, apoptosis occurs (programmed cell death)
what happens during the replication checkpoint
ATM/ATR kinases halt the cell if issues are detected. cell verifies that all dna has been successfully replicated, replication errors or incomplete replication stalls progression to mitosis so repairs can be made before entering
what happens during spindle attachment checkpoint
ensures proper chromosomal alignment and attachment of fibers
- mitotic checkpoint complex (MCC) monitors whether all chromosomes are correctly attached to spindle fibers via kinetochore, MCC inhibits anaphase-promoting complex/cyclosome (APC/C) thus delaying anaphase until all chromosomes are properly aligned at the metaphase plate to prevent unequal chromosomal segregation (aneuploidy)
what is the process of chromatid separation? what does this entail?
- preparation during prophase: chromsomes (made of two sister chromatids connected at a centromere) condense, become visible. mitotic spindle forms, comprimise MTs that extend from entrosome towards chromosomes
- alignment at metaphase: chromosomes align at metaphase plate with centromeres connected to spindle fibers, alignment ensured at checkpoint
- activation at anaphase: once checkpoint statisfied, separase cleaves cohesin and allows them to separate
- chromatid movement: spindle fibers shorten, pulling each one towards opposite poles of cells, these become individual chromosomes
- telophase: chromsomes decondense at poles, nucelear envelope reforms, prepares for cytokinesis
what proteins are involved in chromatid separation? what are their functions?
cohesin: holds sister chromatids together until anaphase
separase: cleave cohesin and allows for separation
spindle checkpoint protein: ensures proper attachment of chromosomes to spindle before anaphase
what is apoptosis?
programmed cell death
what are the structural changes during apoptosis
- chromatin condenses (compact, dense), shrinking the cytoplasm
- nucleus becomes fragmented, DNA laddering (DNA fragmentation leaving bands of dna that look like a ladder), membrane blebs (forms small apoptotic bodies, visual looks like its sounds bleb bleb bleb squiggley), cell fragmentation (breaks to smaller piece containing fragmented organelles and dna)
- phagocytosis occurs (phagocytic cells recognize and engulf apoptotic bodies to prevent inflammation)
- apoptotic body is absorbed by a phagocytic cell (cell breaks down and digests apoptotic bodies)
what are the components of the mitotic spindle?
- microtubules (astral - project outwards, kinteochore - attach to chromosomes, overlap - link to the poles)
- motor proteins (kesin-related (+) - towards plus end poles pushing poles apart, dynein (-) - towards minus end pulling chromsomes toward centrosome)
- chromosomes (chromatids - seperate durig mitosis)
- centrosome (centrioles - organize microtubules and pcm - material around centriole)
what is poleward flux and its components
movement of MTs and associated structures toward spindle poles during mitosis
after MTs shorten and disassemble, motor proteins facilitate movement, dynein moves towards end of MTs, driven by depolymerization of MT ends at + side, pulling tubulin subunits towards opposite poles. this process helps shorten kinteochore MTs after chromosomal alignment at metaphase plate, allowing correct positioning
what are the 2 separation forces?
- MT disassembly drives chromatid movement (Ndc80 complex and kinteochore): Ndc80 complex stabilizes kinetochore-MT attachment, ensuring disassembly occurs at kinetochore, generates force for chromatid movement. kinetochores cpature MTs and promote disassembly directly at sites which pulls chromatids towards spindle poles (leads to disassembly and shortening at kinetochore)
- poleward flux at onset of anaphase: more general, refers only to movement of MTs towards spindle poles cus of depolymerization of MTs at plus ends, results in overall shortening of MT across spindle
what is translocation across the outer membrane? explain the steps and process
definition: movement of proteins and molecules from cytosol into intermembrane space of mitochondria
- targeting signal: proteins moving to there have signal sequence that directs them; receptors recognize it through translocator protein on the outer membrane
- translocator complex: TOM (translocase of the outer membrane) complex binds to protein to be translocated and helps pull thru, signal sequeunce interacts with TOM, facilitates insertion of protein into outer membrane
- protein integration: protein unfolded to fit through translocator channel (usingg energy from ATP), which is then threaded through TOM and into intermembrane space
- release and destination: once across OM, protein may be further processed and transported by other translocator complexes
how are ribosomes directed to the ER membrane?
- recognition: synthesized proteins have a signal sequence at the n-terminus so they can be recognized by binding to the signal recognition particle (SRP), stopping translation and leading the SRP to the ER membrane
- targeting: SRP ribosome complex binds to SRP receptor on ER membrane, the ribosome is transferred to a (translocon) protein channel on ER membrane and inserted into it, allowing translaton to resume and the protein to enter the ER lumen
- release: signal peptide is cleaved off by signal peptidase, ribosome finishes translation and dissociates from translocon
- recycling: SRP and SRP receptor recycling is released and recycled back to cytosol, SRP receptor returns to membrane and receives another SRP ribosome complex, ribosome detaches and continues to synthesize other proteins in the cytoplasm
what are SNARE proteins and their subtypes
SNARE: Soluble NSF Attachmet Protein REceptor
membrane-associated proteins involved in vesicle trafficking and membrane fusion
- v-SNARE (vesicle): on membrane of transport vesicles, proteins recognize and bind to cargo-specific molecules, ensure vesicles are targeted correctly
- t-SNARE (target): on target membrane, bind to v-SNAREs to facilitate vesicle docking and fusion
what are trans-SNARE complexes
t-snare + v-snare make this complex: coiled-coil complex ensuring specific fusion of vesicle with target membrane, allowing delivery of vesicle’s contents to correct location (driven by energy released when forming trans-SNARE complex)
what are the 2 models for protein transport between cisternae
- cisternal maturation model - dynamic: cisternae mature from cis face to trans face (entry to exit of Golgi) cisterna change themselves over time
- newly formed cis cisternae mature as they move through the Golgi and acquire enzymes, modifying proteins as they move, as they matural, they go from cis > medial > trans cisternae, fuzing with the next, proteins are modified without moving cisternae between them. - vesicular transport model - static:
proteins are moved from one cisterna to the next via transport vesicles discrete veiscles move contents from one cisterna into another, not maturing
- proteins are packed into vesicles at one cisterna, bud off and fuse with the next to transfer contents, each retains its identity throughout transport process and vesicles carry specific enzymes required for process
what are the 3 types of membrane proteins
- integral: moves molecules across membrane, across membrane, contain channels and carriers to facilitate selective transport (include ion channels, pumps, receptors)
- peripheral: attaches to the membrane’s inner and outer surface temporarily, doesn’t embed across the lipid bilayer, interacts with integral membrane proteins and phospholipids (i.e. spectrin, ankyrin)
- lipid-anchored: embeds in the lipid bilayer via a covalently bonded lipid molecule (i.e. GPCRs, prenyl, fatty-acid anchored)
explain the structure and function of Na+-K+ ATPase
P-type ATPase pump, that relies on phosphorylation of an intracellular aspartic acid residue (amino acid) and ATP hydrolysis to establish and maintain sodium and potassium gradients across the plasma membrane (consumes 2/3 of cell’s ATP, contributing to membrane potential by exporting 3 Na+ ions out for every 2 K+ in)
explain KcsA K+ channels and their structure
used for selective passage of potassium ions across cell membrane to contibute to the generation of a cell membrane potential
composed of 4 subunits, each with 2 transmembrane helices (M2 helix and M1 helix), where M2 is responsible for controlling the gating mechanism, has a pore-forming segment made of a helix and non-helical loop to form a selectivity filter (which allows only potassium ions to pass through, excluding others and M2 controls opening and closing of the channel)
explain the structure and function of a selectivity filter
made of 5 rings within a pore, each with a diameter of 3Å (0.3 nanometres), closely corresponding to the size of K+ ions (2.7Å or 0.27 nanometres)
- lined with oxygen atoms to interact with K+ ions, but not smaller molecules (when K+ interacts, it loses its hydration cell, interacts with oxygen all the way through, allowing it to flow rapidly)
explain the structure and function of glutamate receptors
glutamate activates both ionotropic and metabotropic receptors, both are permeable to Na+ and K+, but NMDA receptors also require glycine for activiation which are also permeable to Ca2+ (req. for intracellular signalling). they are regulated by factors like voltage-dependent Mg2+ to block channels at resting to prevent Ca2+ influx, and Mg2+ expelling to allow receptors to open and permit Ca2+ entry upon membrane depolarization
This chain of events leads to NMDA receptors integrating signals from presynaptic glutamate release and postsynaptic membrane potential changes
what are the types of receptors and how do they work
- ionotropic receptors: multiple subunits that form ion channels, directly bind to neurotransmitters and allow ions to cross the cell membrane, alter the membrane potential and produce rapid-immediate short-term responses
- metabotropic receptors: protein with 7 transmembrane domains (GPCRs), don’t form channels, but instead activate intracellular signaling cascades by binding to neurotransmitters to produce slow, long-term signals (control ion channel activity via second messengers)
what is long-term potential and how is it formed?
underlies learning and memory, strengthens synaptic connections between neurons.
- presynaptic activation: a high frequecy stimulus leads to sustained release of glutamate which then binds to NMDA and non-NMDA glutamate receptors
- postsynaptic activation: that binding allows for an influx of Na+, and opening of NMDA receptors, but Mg2+ blocks Ca2+ from entering depolarizing the post-synaptic membrane
- NMDA receptor activation: if depolarization is sufficient, Mg2+ releases its block on NMDA and allows Ca2+ to enter, the influx triggers a second message and the intracellular signalling cascades
- the potential is brought back to pre-synaptic potential and non-NMDA receptor expression is increased
what is the structure of g-proteins?
composed of 3 subunits (alpha, beta, gamma), alpha subunit acts as the binding site and is responsible for signalling, beta and gamma subunits are anchored to the plasma membrane and stabilize the complex
how do g-proteins function? what is their role?
gtp binding proteins, serve as molecular switches that relay signals from GPCRs to target ion channels or enzymes
- ligand binds to GPCR on cell membrane
- activates GPCR and causes it to interact with the g-protein
- alpha subunit phorphorylates GDP –> GTP, activating the g-protein
- activated alpha subunit, with beta-gamma complex, modules target proteins
- deactivated by hydrolyzing GTP –> GDP to turn off the signal and reassemble the original complex
how does vision involve GPCRs? which one/ones?
involves rhodopsin
1. one rhodopsin molecule absorbs one photon
2. five hundred transducin molecules are activated
3. 500 phophodiesterase molecules are activated
4. 10^5 cyclic GMP molecules are hydrolyzed
5. 250 Na+ channels close
6. 10^6-10^7 Na+ ions per second are prevented from entering the cell in ~1 second
7, rod cell membrane is hyperpolarized by 1 mV
