Exam One

Protein Sorting

proteins are selectively transferred from the cytosol where they are made to the compartment where they will be used

What organelles obtain their protein directly from the cytosol?

mitochondria

chloroplasts

interior of nucleus

What organelles obtain their proteins and lipids from the endoplasmic reticulum?

Golgi apparatus

lysosomes

endosomes

inner nuclear membrane

What are the three mechanisms organelles use to import proteins?

through pores within the membrane of the nucleus

through protein translocators in the ER, mitochondria, or chloroplasts (proteins unfold and thread through)

by vesicles between organelles within the endomembrane system (pinch off from one organelle and fuse with another)

Günter Blobel

1999 Nobel Prize in Physiology and Medicine for the discovery of protein signals that govern their transport and localization in the cell

Signal Sequences

direct proteins to their correct organelle and are required and sufficient for protein targeting

typically 15-60 amino acids long

often removed from protein after sorting

can be in the middle of a protein

Where do proteins without a signal sequence go?

remain in cytosol

GFP-Signal Peptide Fusion Proteins

can report on targeted movement of proteins

Endoplasmic Reticulum (ER)

specializes in manufacturing

proteins destine for the ER, Golgi, lysosome, peroxisome, endosome, plasma membrane, and secretion must first go here

Rough Endoplasmic Reticulum

studded with ribosomes for protein synthesis

Signal-Recognition Particle (SRP)

binds to the signal sequence of the protein being made by the ribosome

receptor binds and recruits ribosome to ER

protein is threaded through a translocator and into the ER as translation takes place

Signal Peptidase

removes the signal sequence

Stop-Transfer Sequence

stops the transfer of the protein chain

hydrophobic and remains in the membrane as a transmembrane domain

Endomembrane System

the interconnected network of membrane enclosed organelles

ER, Golgi, lysosomes, peroxisomes, endosomes

Vesicles

bud off of and fuse with membranes for protein and lipid trafficking

Clathrin Proteins

assemble into a basket-like network around vesicle

Adaptins

link cargo receptors to clathrin network

Coat Proteins

COPI and COPII

coat vesicles

Cytoskeleton

helps position organelles within the cell

vesicles transported along the tracks

Microtubule Motor Proteins

can carry vesicles as they walk along the microtubule

kinesin and dynein

Rab Proteins

in combination provide vesicles with unique markings for the target membrane

Tethering Proteins

bind to Rab proteins

SNARE Proteins

firmly dock the vesicle to the membrane

catalyze the fusion of the two membranes

two proteins, v (on the vesicle) and t (on the target membrane), wind together

Nuclear Pores

made up of over 30 individual proteins that contain a tangled meshwork that prevents large molecule movement but allow small soluble molecules to diffuse through

where materials move in and out of the nucleus

Nuclear Localization Signal (NLS)

found on proteins transported to the nucleus

bind to the nuclear import receptor

Mitochondrial Signal Sequence

recognized by a receptor on the outer membrane until translocator takes over then recognized by translocator on inner membrane

Stem Cell

a cell that retains the ability to divide and recreate itself (self-renewal) while also having the power (potency) to generate progeny capable of specializing to a more differentiated cell type

Hematopoietic Stem Cells (HSC)

stem cells to replace blood cells

Niche

the microenvironment that surround stem cells and regulates them

keeps stem cells uncommitted until the cell leaves the environment

cell-to-cell interaction, cell signaling, cytoplasmic determinants, transcription factors, and epigenetic regulation

Hematopoietic Stem Cell Niche

adjacent to osteoblasts that line bone cavity (endosteal) and cells that line blood vessels (perivascular)

Totipotent Stem Cells

stem cells that can become all cell types

Pluripotent Stem Cells

stem cells that can become all cells of an embryo

Multipotent Stem Cells

stem cells that are restricted by tissue

Embryonic Stem Cells (ESC)

isolated from a human blastocyst

cell can be forced to a specific fate and transplanted into patient

Somatic Cell Nuclear Transfer (SCNT)

haploid nucleus removed from oocyte and replaced with nucleus from somatic cell

the embryonic stem cells are then removed from the inner cell mass

makes patient specific stem cells and to clone

What are the problems with using human embryonic stem cells (hESC)?

potential immunogenicity

ethical issues regarding the use of human embryos

Induced Pluripotent Stem Cells (iPSC)

differentiated somatic cells that are reprogrammed to stem cells with the expression of three to four factors

What are the issues of using iPSCs in humans?

tumorgenicity

immunogenicity

heterogeneity

Neoblasts

planarian stem cells

the only mitotic cell population in planarians

Epimorphosis

the de novo rebuilding of anatomy

Blastema

mass of cells capable of growth and regeneration into organs or body parts

How do neoblasts know what to do?

gradient system

Wnt for anterior/posterior

BMP for dorsal/ventral and medial/lateral

Positional Control Genes (PCGs)

genes that control the location of body parts and tissues during regeneration

expressed in muscle cells which are required for planarian planning

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)

repetitive DNA sequences with spacers in between

CRISPR-Associated Genes (Cas)

genes that encode proteins to cut foreign DNA and RNA

Type 1 Expression & Interference

multiple Cas proteins in a complex with crRNA

CRISPR RNA (crRNA)

guides CRISPR protein to destroy a sequence of DNA

Type 2 Expression & Interference

a single Cas protein with a crRNA and a tracrRNA

Protospacer Adjacent Motif (PAM)

the binding signal for Cas9

target sequence must be immediately adjacent to it

not present in bacterial genome

Non-Homologous End Joining (NHEJ)

repairs a double stranded DNA break without a template

error prone

results in insertions and deletions

Homology-Directed Repair (HDR)

repair a double stranded DNA break with a template

error free

allows for the insertion of a point mutation or DNA fragment

CRISPR Interference (CRISPRi)

dCas9 fused to transcriptional repressor proteins and represses expression of specific genes

Cas9 nuclease function removed but can still bind to target DNA sequences

What are the ethical concerns of DNA editing?

safety

informed consent

justice and equity

moral and religious