Luigi Karlo M. AlladoBio - 4

• The biosynthetic pathway of a eukaryotic cell consists of a series

of distinct membrane-bound organelles

• Materials are carried between compartments by vesicle that bud from donor membranes and fuse with acceptor membranes

These electron micrographs show the membranes of these vesicles to be covered on their outer (cytosolic) surface by a distinct protein coat

a protein coat formed from soluble proteins that assemble on the cytosolic surface of the donor membrane at sites where budding takes place

Activation of a small G protein recruited to the site

Act as mechanical device that causes membrane to curve and form budding vesicle

Provide a mechanism in selecting the components to be carried by vesicle

cargo consisting of secretory, lysosomal, and membrane proteins to be transported

The machinery required to target and dock the vesicle to the correct acceptor membrane

move materials from the ER “forward” to the ERGIC and Golgi complex.

Contains a number of proteins that were first identified in mutant yeast cells that were unable to carry out transport from the ER to the Golgi complex

select and concentrate certain components for transport in vesicles

enzymes that act at later stages in the biosynthetic pathway, such as the glycosyltransferases of the Golgi complex

membrane proteins involved in the docking and fusion of the vesicle with the target compartment

Membrane proteins that are able to bind soluble cargo

Mutations in one of these cargo receptors has been linked to an inherited bleeding disorder

a small G protein called Sar1, which is recruited specifically to the ER membrane

Sar1 plays a regulatory role

Sar1-GDP molecules have been recruited to the ER membrane by a protein called a GEF (guanine-exchange factor) that catalyzes the exchange of the bound GDP with a bound GTP

each Sar1-GTP molecule has extended a finger-like helix along the membrane within the cytosolic leaflet. This event induces the curvature of the lipid bilayer at that site

a dimer composed of two COPII polypeptides(Sec23 and Sec24) has been recruited by the bound Sar1-GTP.

Transmembrane cargo accumulates within the forming COPII vesicle as their cytosolic tails bind to the Sec24 polypeptide of the COPII coat

the remaining COPII polypeptides (Sec13 and Sec31) have joined the complex to form an outer structural scaffold of the coat.

Two copies of the Sar1-Sec23-Sec24 complex (shown in red, magenta, and blue, respectively) that would form the inner layer of the COPII

move materials in a retrograde direction from the ERGIC and Golgi stack

“backward” toward the ER and from trans Golgi cisternae

“backward” to cis Golgi cisternae first identified in experiments in which

cells were treated with molecules similar in structure to GTP

Unlike GTP, cannot be hydrolyzed

accumulate in the presence of a nonhydrolyzable GTP analogue

the coat contains a small GTP binding protein, called ARF1, whose bound GTP must be hydrolyzed before the coat can disassemble

proteins are maintained in an organelle by a combination of two mechanisms:

Retention - may be based primarily on the physical properties of the protein ex. Soluble proteins that are part of large complexes or membrane proteins with short transmembrane domains are not likely to enter a transport vesicle.

Retrieval of “escaped” molecules back to the compartment in which they normally reside

trans Golgi network Major sorting station

Sorting and transport of lysosomal enzymes

Lysosomal proteins are synthesized on membrane-bound ribosomes of the ER and carried to the Golgi complex along with other types of proteins

soluble lysosomal enzymes are specifically recognized by enzymes that catalyze the two-step addition of a phosphate group to certain mannose sugars of the N-linked carbohydrate chains

Lysosomal enzymes are transported from the TGN in clathrin-coated vesicles

Lysosomal enzymes are escorted from the TGN by a family of adaptor proteins called GGAs

The outer ends of the GGA adaptors bind to clathrin molecules

GGA adaptors bind to a sorting signal in the cytosolic tails of the mannose 6-phosphate receptors

MPRs in the TGN membrane and lysosomal enzymes within the TGN lumen become concentrated into clathrin-coated vesicles

production of clathrin-coated vesicles begins with the recruitment to the membrane of a small GTP-binding protein

Lysosomal proteins are not the only materials that are exported from the TGN

membrane proteins destined for the plasma membrane and secretory materials destined for export from the cell are also transported from the TGN, but the mechanisms are poorly understood

requires specific interactions between different membranes

Selective fusion is one of the factors that ensures a directed flow through the membranous compartments of the cell

1. Movement of the vesicle toward the specific target compartment

2. Tethering vesicles to the target compartment

3. Docking vesicles to the target compartment

4. Fusion between vesicle and target membranes

fusion of a secretory vesicle or secretory granule with the plasma membrane and subsequent discharge of its contents

continual basis in most cells, as proteins and other materials are delivered to both the plasma membrane and extracellular space

membrane fusion produces an opening through which the contents of the vesicle or granule are released into the extracellular space

an animal cell’s digestive organelles contains at least 50 different hydrolytic

enzyme sproduced in the rough ER and targeted to these organelles

The enzymes of a lysosome share an important property: allhave their optimal activity at an acid pH and thus are acid hydrolases.

Optimum pH is 4.6 breakdown of materials brought into the

cell from the extracellular environment

Lysosomes also play a key role in organelle turnover, that is, the regulated destruction of the cell’s own organelles and their replacement

Process is called autophagy Once the digestive process in the

autophagolysosome has been completed, the organelle is termed a residual body

90% in a plant cell Solutes and macromolecules are temporarily

stored May also store host of toxic compounds tonoplast,contains a number of active transport

systems that pump ions into the vacuolar compartment to a concentration much higher than that in the cytoplasm or the extracellular fluid

Osmosis sites of intracellular digestion have some of the same acid hydrolases found in

lysosomes

Endocytosis - is primarily a process by which the cell internalizes cell-surface receptors and bound extracellular ligands

bulk-phase endocytosis - is the nonspecific uptake of extracellular fluids

removes portions of the plasma membrane and may function primarily in the recycling of membrane between the cellsurface and interior compartments

in contrast, brings about the uptake of specific extracellular macromolecules (ligands) following their binding to receptors on the external surface of the plasma membrane

provides a means for the selective and efficient uptake of macromolecules that may be present at relatively low concentrations in the extracellula fluid

Cell eating In most animals, phagocytosis is a

protective mechanism rather than a mode of feeding

Not all bacteria ingested by phagocytic cells are destroyed.

some species hijack the phagocytic machinery

to promote their own survival in the body

Have 2 subcompartments in which imported proteins take place: boundary membrane and the internal matrix

Proteins destined for a peroxisome possess a peroxisomal targeting signal, either a PTS for a peroxisomal matrix protein or an mPTS for a peroxisomal membrane protein

PTS receptors bind to peroxisome-destined proteins in the cytosol and shuttle them to the surface of the peroxisome

The PTS receptor apparently accompanies the peroxisomal protein through the boundary membrane into the matrix and then recycles back to the cytosol to escort another protein

Unlike mitochondria and chloroplasts, whose imported proteins must assume an unfolded state, peroxisomes are somehow able to import peroxisomal matrix proteins in their native, folded conformation, even those that consist of several subunits. The mechanism by which peroxisomes are able to accomplish this daunting assignment remains a matter of speculation.

have four subcompartments into which proteins can be delivered

an outer mitochondrial membrane (OMM),

inner mitochondrial membrane (IMM), intermembrane space,

and matrix

Chloroplasts have six subcompartments into which proteins can be delivered: an inner and outer envelope membrane and

intervening intermembrane space, as well as the stroma, thylakoid membrane, and thylakoid lumen

Chloroplast and mitochondrial import mechanisms exhibit many similarities, although their translocation machineries have evolved independently

1. the vast majority of chloroplast proteins are imported from the cytosol, 2. the outer and inner envelope membranes contain

distinct translocation complexes (Toc and Tic complexes,

respectively) that work together during import, 3. chaperones aid in the unfolding of the polypeptides in

the cytosol and folding of the proteins in the chloroplast,

and 4. most proteins destined for the chloroplast are

synthesized with a removable N-terminal sequence (termed the

transit peptide).