Intracellular compartments and protein sorting

Introduction

Different proteins need to be sent to different parts of a eukaryotic cell, or, in some cases, exported out of

the cell and into the extracellular space. How do the right proteins get to the right places?

Cells have various shipping systems, kind of like molecular versions of the postal service, to make sure

that proteins arrive at their correct destinations. In these systems, molecular labels (often, amino acid

sequences) are used to "address" proteins for delivery to specific locations. Let’s take a look at how these

shipping systems work.

1) The nucleus and nuclear pore complex

2) ER, golgi, endosomes and lysosomes
3) mitochondria

Overview of cellular shipping routes

Translation of all proteins in a eukaryotic cell begins in the cytosol (except for a few proteins made in

mitochondria and chloroplasts). As a protein is made, it passes step by step through a shipping "decision

tree." At each stage, the protein is checked for molecular tags to see if it needs to be re-routed to a

different pathway or destination.

 Intracellular compartments and protein sorting

Mechanisms of protein transport

 Transport through nuclear pores ( gated transport)

 Transport across membranes
 Transport by vesicles

Nuclear pore complex and envelope

 Contains the genome, site of DNA replication and site of transcription

 Large multi-protein assembly that forms a channel through the nuclear envelope
 composed of 30 proteins called nucleoporins
 Nucleoporins regulate transport in and out of pores
 FG(phenylarine -glycine) forms a long hydrophobic sieve that blocks diffusion of larger
molecules

Nuclear lamina: regulates the size, shape and chromatin patterns of the nucleus

Nucleolus: site of ribosome assembly and rRNA synthesis, larger and more numerous in cells actively
synthesizing proteins and rapidly growing cells ( embryonic cells, malignant tumour cells)

Cajal bodies: snRNPs and snoRNPs maturation, snRNP recyclying and resetting

Granule cluster: snRNPs and RNA processing components storage

 Intracellular compartments and protein sorting

Interchromatin: speed up RNA processing

Mechanism of protein transport into the nucleus:

*Free diffusion for small molecules

 Aim :To get Nuclear proteins across

nuclear pore
 Require importins as receptors e.g
importin B, importin a, NLS protein
(nuclear localization signal) which exist
as dimers
1) Proteins with NLS stretch bind
to importin. Importin cargo
complex enters through the
NPC
2) The composition of the npc
changes as protein passes
through. Release of proteins
from the importins, once
entered Ran-GTPases help
recycle receptors
3) Importin-Ran-GTP complex are
exported. Exportins export
substances out of the NPC and
contain nuclear export signals.

Questions:
1) What is the function of importins and exportins?
2) What are required to transfer cargo into the NPC?
3) What is the role of Ran-GTP in the process?
 Intracellular compartments and protein sorting

Uptake of proteins by the mitochondrion

 Double membrane bound organelle

 Specialize in ATP synthesis
 Most of its proteins are encoded in the nucleus and imported
from the cytosol

Mechanism:
1) Proteins must be unfolded first by chaperones and require a
mitochondrial sequence
2) Outer mitochondrial includes a protein import complex known as the TOM
complex (transport onto membrane) which includes a receptor and a channel
3) Proteins destined for the inner membrane into the matrix require the TIM
complex (Transport into Matrix). The Tim 22 mediates the insertion of subset of
membrane proteins. The tim 23 translocate soluble proteins.
4) Movement into the matrix is voltage dependent
5) The Sam complex helps to translocate and fold outer membrane proteins**
6) Once inside the matrix, polypeptides get their signals cleaved by peptidases.

Questions
1)
 Intracellular compartments and protein sorting

Import of protein to the ER

The signal peptide that sends a protein into the endoplasmic reticulum during translation is a series of

hydrophobic (“water-fearing”) amino acids, usually found near the beginning (N-terminus) of the protein.

When this sequence sticks out of the ribosome, it’s recognized by a protein complex called the signal-

recognition particle (SRP), which takes the ribosome to the ER. There, the ribosome feeds its amino acid

chain into the ER lumen (interior

Rough ER

The rough endoplasmic reticulum (rough ER) gets its name from the bumpy ribosomes attached to its

cytoplasmic surface. As these ribosomes make proteins, they feed the newly forming protein chains into

the lumen. Some are transferred fully into the ER and float inside, while others are anchored in the

membrane.

 Protein synthesis

 Integral

membrane

proteins

 Lysosomal

enzymes

 Core glycosylation

Since the rough ER helps modify proteins that will be secreted from the cell, cells whose job is to secrete

large amounts of enzymes or other proteins, such as liver cells, have lots of rough ER.

Smooth ER

continuous with the rough ER but has few or no ribosomes on its cytoplasmic surface. Functions of the

smooth ER include: Synthesis of carbohydrates, lipids, and steroid hormones, Detoxification of

medications and poisons,Storage of calcium ions

Intracellular compartments and protein sorting
 Intracellular compartments and protein sorting

 Stop transfer signal

 Internal shift transfer

 Start and stop internal

signal sequence
 Can be restarted and have
multiple membrane
proteins

Tail anchored proteins

 Post-translational pathway
 Membrane proteins (tail anchored)
 C-terminal -> hydrophobic a helix
 Recognition from the SRP not possible

Questions:
1) What is the function of the rough ER?
2) What is the function of the smooth ER?
3) Describe the mechanism of protein transport into the ER
 Intracellular compartments and protein sorting

Peroxisomes

Single membrane, no DNA or ribosomes. All their proteins are coded in the nucleus.
They import proteins from the cytosol. It houses enzymes involved in oxidation
reactions, which produce hydrogen peroxide as a by-product. The enzymes break down
fatty acids and amino acids, and they also detoxify some substances that enter the body.
For example, alcohol is detoxified by peroxisomes found in liver cells.

Importantly, peroxisomes—unlike lysosomes—are not part of the endomembrane

system. That means they don't receive vesicles from the Golgi apparatus.

Proteins needed in the peroxisome have a specific sequence of amino acids called

a peroxisomal targeting signal. The classic signal consists of just three amino acids, serine-

lysine-leucine, found at the very end (C-terminus) of a protein. This pattern of amino acids is

recognized by a helper protein in the cytosol, which brings the protein to the peroxisome

Mitochondrial, chloroplast, and nuclear targeting are generally like peroxisomal targeting. That

is, a certain amino acid sequence sends the protein to its target organelle (or a compartment

inside that organelle). However, the nature of the "address labels" is different in each case.

Peroxins*ATP hydrolysis*

Questions: what is the function of a peroxisome?

2) What is the difference between a peroxisome and a lysosome?
Intracellular compartments and protein sorting