Cellular Biology I - Endomembrane System (PDF)

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It is allowed the use of the material for private and study use, however not for profit and without commercial purposes. Biomedical Sciences I Cellular Biology I Lorena Di Pietro [email protected] The Endomembrane System Endomembrane System IN ALL EUKARYOTIC CELLS DISTINCT COMPARTMENTS, BOUNDED BY MEMBRANE, FORMING AN EXTENSIVE NETWORK OF INTERCONNECTED TUBULES AND FLATTENED SACKS Endomembrane System INCLUDES: - Rough endoplasmic reticulum - Smooth endoplasmic reticulum - Golgi apparatus - Lysosomes - Endosomes Endomembrane System THE ORGANELLES OF THE ENDOMEMBRANE SYSTEM ARE PART OF A DYNAMIC NETWORK IN WHICH MATERIALS ARE SHUTTLED BACK AND FORTH FROM ONE PART OF THE CELL TO ANOTHER Endomembrane System THE MATERIALS ARE SHUTTLED BETWEEN ORGANELLES IN SMALL MEMBRANE-BOUNDED TRANSPORT VESICLES TRANSPORT VESICLES MOVE IN A DIRECTED MANNER Motor proteins and cytoskeletal elements play key roles in the movements of transport vesicles Endomembrane System – FUNCTIONS 1. BIOSYNTHETIC PATHWAY –> PROTEINS ARE SYNTHETIZED IN THE ER, MODIFIED DURING PASSAGE THROUGH THE GOLGI, AND TRANSPORTED FROM THE GOLGI TO VARIOUS DESTINATIONS - (membrane proteins, lysosomal enzymes) 2. SECRETORY PATHWAY –> PROTEINS ARE DESTINATED TO BE SECRETED FROM THE CELL - (extracellular matrix proteins, hormones, neurotransmitters) 3. ENDOCYTIC PATHWAY –> MATERIALS MOVE FROM THE CELL SURFACE TO THE INTRACELLULAR SPACE The Endoplasmic Reticulum (ER) THE WIDEST SUBCELLULAR STRUCTURE IN EUKARYOTIC CELLS NETWORK OF MEMBRANOUS TUBULES AND SACS CALLED CISTERNAE ENCLOSED WITHIN THE ER IS AN EXTENSIVE SPACE (LUMEN) THAT IS SEPARATED FROM THE CYTOSOL BY THE ER MEMBRANE Protein disulfide isomerase (PDI) The Endoplasmic Reticulum (ER) THE OUTER NUCLEAR MEMBRANE IS CONTINUOUS WITH THE ER MEMBRANE, HENCE THE PERINUCLEAR CAVITY IS CONTINUOUS WITH THE ER LUMEN PLASTIC AND DYNAMIC IN SHAPE AND SIZE Protein disulfide isomerase (PDI) The Endoplasmic Reticulum (ER) THERE ARE TWO DISTINCT REGIONS OF ER; EVEN THOUGH THEY ARE CONTINUOUS WITH EACH OTHER, THEY DIFFER IN STRUCTURE AND FUNCTION. - Rough Endoplasmic Reticulum (RER) - Smooth Endoplasmic Reticulum (SER) RER SER Rough Endoplasmic Reticulum (RER) CONTINUOUS WITH THE OUTER NUCLEAR MEMBRANE RIBOSOMES ARE ATTACHED ON THE CYTOPLASMIC SURFACE OF THE MEMBRANE OF RER –> IT APPEARS ROUGH THROUGH ELECTRON MICROSCOPE Rough Endoplasmic Reticulum - FUNCTIONS Synthesis, folding, start of glycosylation, quality control for proteins  Protein synthesis PROTEINS DESTINED TO CYTOSOL, NUCLEUS, MITOCHONDRIA, AND PEROXISOMES, COMPLETE THE PROCESS IN THE CYTOSOL (FREE RIBOSOMES) PROTEINS DESTINED TO ER, GOLGI, LYSOSOMES, PLASMA MEMBRANES, OR TO SECRETION, COMPLETE THE PROCESS IN RER (BOUND RIBOSOMES)  Protein synthesis Proteins must be delivered to the cellular compartment in which they function  Protein maturation PROTEINS ARE FORWARDED TO THEIR “CORRECT ADDRESS” THANKS TO SPECIFIC SORTING SIGNALS, ENCODED BY THE AMINOACIDIC SEQUENCE OR IN THE ATTACHED OLIGOSACCHARIDES Chemical modification of newly translated polypeptides is needed to allow the correct sorting and transport Protein maturation THE TYPICAL SORTING SIGNAL ON A PROTEIN IS A CONTINUOUS STRETCH OF AMINO ACID SEQUENCE (15-60 AA OFTEN FOUND AT THE N-TERMINUS) THE SORTING SIGNALS ARE RECOGNIZED BY SPECIFIC RECEPTORS THAT RESIDE IN THE MEMBRANES OR SURFACE COATS OF BUDDING VESICLES THE SIGNAL SEQUENCE IS OFTEN (BUT NOT ALWAYS) REMOVED FROM THE PROTEIN ONCE IT HAS BEEN SORTED BY SPECIALIZED SIGNAL PEPTIDASES Protein maturation THERE ARE 3 DIFFERENT WAYS BY WHICH PROTEIN MOVES FROM ONE COMPARTMENT TO ANOTHER: 1. gated transport (nuclear pore complexes) 2. protein translocation (throughout protein translocators) 3. vesicular transport Protein maturation Rough Endoplasmic Reticulum - FUNCTIONS SYNTHESIS ONE-THIRD OF THE PROTEINS IN MAMMALS ARE SYNTHESIZED ON RIBOSOMES ATTACHED TO THE RER AND RELEASED INTO THE LUMEN (CO‐TRANSLATIONAL TRANSLOCATION) - secreted proteins - integral membrane proteins - soluble proteins that reside within compartments of the endomembrane system Rough Endoplasmic Reticulum - FUNCTIONS THE PROTEINS THAT ENTER IN THE RER ARE OF TWO TYPES: - water-soluble proteins, which are fully translocated across the ER membrane and are released into the ER lumen - transmembrane proteins, which only partly translocated across the ER membrane and become embedded in it - Some of the transmembrane proteins are destined to reside in the plasma membrane or the membrane of another organelle  Rough Endoplasmic Reticulum - FUNCTIONS The interaction Polypeptides addressed to The signal is recognized between the SRP and RER contain a by a signal recognition the SRP receptor on the hydrophobic signal particle (SRP), that ER membrane is sequence (usually located binds to both the signal followed by the bond at or near the N-terminus) and the ribosome of the ribosome with the translocon SYNTHESIS  Rough Endoplasmic Reticulum - FUNCTIONS The contact of the signal sequence with the interior The nascent polypeptide Upon termination of of the translocon leads to passes across the translation the plug is the displacement of the translocon to the ER reinserted into the plug, that usually seal the lumen translocon channel channel SYNTHESIS Rough Endoplasmic Reticulum - FUNCTIONS The water-soluble proteins are destined either for the lumen of an organelle, including ER, or for secretion SYNTHESIS Rough Endoplasmic Reticulum - FUNCTIONS Integral membrane proteins The translocon orients the contain one or more nascent polypeptide --> hydrophobic transmembrane the N-terminus facing segment that are shunted either the cytosol or the directly from the channel of the lumen of the ER translocon into the lipidic bilayer SYNTHESIS Rough Endoplasmic Reticulum - FUNCTIONS Some of the transmembrane proteins function in the ER, but many are destined to reside in the plasma membrane or the membrane of another organelle SYNTHESIS Rough Endoplasmic Reticulum - FUNCTIONS GLYCOSYLATION MANY OF THE PROTEINS THAT ENTER THE ER ARE CONVERTED TO GLYCOPROTEINS (COVALENT ATTACHMENT OF A SHORT, BRANCHED OLIGOSACCHARIDE SIDE CHAIN) VERY FEW PROTEINS IN THE CYTOSOL ARE GLYCOSYLATED, AND THOSE THAT ARE HAVE ONLY A SINGLE SUGAR ATTACHED TO THEM OLIGOSACCHARYLTRANSFERASE ADDS CARBOHYDRATES TO THE NASCENT PROTEINS Rough Endoplasmic Reticulum - FUNCTIONS GLYCOSYLATION ROLES OF GLYCOSYLATION: protects protein from degradation; holds protein in the ER until it is properly folded; help protein to be guided to the right final destination (serves as a sorting signal) SUGAR CHAINS OF GLYCOPROTEINS FORM PART OF THE CELL’S OUTER LAYER CALLED GLYCOCALYX Rough Endoplasmic Reticulum - FUNCTIONS GLYCOSYLATION A PREFORMED PRECURSOR OLIGOSACCHARIDE IS HELD IN THE ER MEMBRANE BY A SPECIAL LIPID MOLECULE CALLED DOLICHOL, AND IT IS TRANSFERRED IN A SINGLE ENZYMATIC STEP Rough Endoplasmic Reticulum - FUNCTIONS GLYCOSYLATION THE OLIGOSACCHARIDE CHAIN IS TRANSFERRED TO THE N- TERMINUS OF THE NASCENT POLYPEPTIDE AND THEN UNDERGOES A PROCESS OF GRADUAL MODIFICATION WHILE STILL IN THE ER, THREE GLUCOSES AND ONE MANNOSE ARE QUICKLY REMOVED FROM THE OLIGOSACCHARIDES OF MOST GLYCOPROTEINS Rough Endoplasmic Reticulum - FUNCTIONS FOLDING AND QUALITY CONTROL THE NEWLY SYNTHETIZED GLYCOPROTEIN IS SCREENED FOR A SYSTEM OF QUALITY CONTROL (MOLECULAR CHAPERONES) THAT DETERMINES WHETHER IT IS FIT TO MOVE ON THE NEXT COMPARTMENT MISFOLDED PROTEINS ARE RECOGNIZED, DISLOCATED TO THE CYTOSOL AND PROCESSED FOR DEGRADATION Congenital diseases of Glycosylation - Inherited metabolic disorders affecting nearly every organ system - Are missing one of the enzymes that is required for glycosylation - Mutations that lead to partial disruption of the glycosylation pathway in the ER Cystic Fibrosis - Is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene, encoding a transport channel of the plasma membrane - The ER quality control targets the most frequent disease-causing mutant protein → even though the mutant protein could function normally as a chloride channel if it reached the plasma membrane, it is retained in the ER and degraded Rough Endoplasmic Reticulum - FUNCTIONS Synthesis, folding, start of glycosylation, quality control for proteins that will be secreted Modifies the composition of membranes Major site of synthesis of new membranes in the cell Rough Endoplasmic Reticulum - FUNCTIONS THE PROTEINS SYNTHETIZED IN THE RER WILL BE INSERTED IN A PREDICTABLE ORIENTATION INTO THE LIPID BILAYER THE CARBOHYDRATE CHAINS ARE ALWAYS PRESENT ON THE INTERNAL SIDE OF VESICLES BECAUSE THEY BECAME THE EXTERNAL SIDE OF THE PLASMA MEMBRANE FOLLOWING THE FUSION OF VESICLES WITH IT Rough Endoplasmic Reticulum - FUNCTIONS The lumen of secretory pathway compartments are very similar to the extracellular space Smooth Endoplasmic Reticulum (SER) LACKS RIBOSOMES USUALLY SMALLER THAN RER FARTHER FROM THE NUCLEUS TUBULAR CAVITIES IN CONTINUOUS WITH RER Smooth Endoplasmic Reticulum - FUNCTIONS Sequestering calcium ions within the cytoplasm of cells Synthesis of steroid hormones in the endocrine cells of the gonad and adrenal cortex Detoxification of drug and harmful compounds in liver cells Principal site of production of lipoprotein particles in liver cells Rough Endoplasmic Reticulum - FUNCTIONS MOST MEMBRANE LIPIDS, INCLUDING BOTH PHOSPHOLIPIDS AND CHOLESTEROL, ARE SYNTHETIZED ENTIRELY WITHIN THE ER, EXCEPT FOR SPHINGOMYELIN AND GLYCOLIPIDS WHOSE SYNTHESIS IS COMPLETED IN THE GOLGI COMPLEX THE ENZYMES INVOLVED IN THE SYNTHESIS OF PHOSPHOLIPIDS ARE INTEGRAL PROTEINS OF THE ER MEMBRANE, THAT HAVE THEIR ACTIVE SITES FACING THE CYTOSOL, WHERE ALL THE REQUIRED METABOLITES ARE FOUND Smooth Endoplasmic Reticulum - FUNCTIONS CONTAINS ENZYMES THAT CATALYZE A SERIES OF REACTIONS TO DETOXIFY BOTH LIPID-SOLUBLE DRUGS AND VARIOUS HARMFUL COMPOUNDS PRODUCED BY METABOLISM CYTOCHROME P450 ENZYME FAMILY → WATER-INSOLUBLE DRUGS OR METABOLITES THAT WOULD OTHERWISE ACCUMULATE TO TOXIC LEVELS ARE RENDERED SUFFICIENTLY WATER-SOLUBLE TO LEAVE THE CELL AND BE EXCRETED IN THE URINE Smooth Endoplasmic Reticulum - FUNCTIONS 1. When large quantities of certain compounds, enter the circulation, detoxification enzymes are synthesized in the liver in unusually large amounts, and the SER doubles in surface area within a few days 2. Once the drug has disappeared, the excess SER membrane is specifically and rapidly removed by a lysosome-dependent process called autophagocytosis  Smooth Endoplasmic Reticulum - FUNCTIONS MUSCLE CELLS HAVE AN ABUNDANT SPECIALIZED SER (SARCOPLASMIC RETICULUM) WHICH SEQUESTERS CA2+ FROM THE CYTOSOL THE RELEASE AND REUPTAKE OF CA2+ TRIGGER THE CONTRACTION AND RELAXATION, RESPECTIVELY, OF THE MYOFIBRILS DURING EACH ROUND OF MUSCLE CONTRACTION The Endoplasmic Reticulum Cells that secrete large RER amount of proteins (pancreas or salivary glands) Different ratios RER/SER in different type Steroid-producing of cells endocrine glands SER Skeletal muscle, kidney tubules, liver Golgi complex PILE OF DISC-SHAPED FLATTENED TANKS (CISTERNAE) WITH THICKENED BORDERS ASSOCIATED TUBULES AND VESICLES FROM FEW TO SEVERAL THOUSANDS/CELL Golgi Complex Golgi complex DYNAMIC STRUCTURE VESICLES BUD FROM A PERIPHERAL TUBULAR DOMAIN OF EACH CISTERNA IT IS A POLARIZED ORGANELLE Golgi complex CIS FACE - Closer to ER - Up-taking incoming vesicles (entry face) TRANS FACE - Opposite side - Releasing secretory vesicles (exit face) Golgi complex VESICLES FROM THE ER FUSE TO FORM THE ER-GOLGI INTERMEDIATE COMPARTMENT (ERGIC)  Golgi complex Sorting station where proteins are segregated to their destination CARGOES ARE SHUTTLED THROUGH THE GOLGI STACK FROM THE CGN TO TGN (vesicular transport model) Sorting station that identifies proteins that can proceed to the next compartment Golgi complex SEVERAL GROUPS OF PROTEINS SEEM TO PLAY AN IMPORTANT ROLE IN: 1) supporting mechanically the membranous element of Golgi; 2) directing the movement of vesicles and tubules that move along the complex (motor proteins); 3) regulating the disassembly and reassembly of the Golgi complex during mitosis Golgi complex - FUNCTIONS HANDLING/PACKAGING CENTRE OF THE CELL Golgi complex - FUNCTIONS Modification and synthesis of the carbohydrate portions of glycoproteins Golgi complex - FUNCTIONS GLYCOSYLATION THE N-LINKED OLIGOSACCHARIDES THAT WERE ADDED TO PROTEINS IN THE ER SERVE TO HELP PROTEINS FOLD AND TO HELP TRANSPORT MISFOLDED PROTEINS TO THE CYTOSOL FOR DEGRADATION (QUALITY CONTROL) THE GOLGI GENERATES THE HETEROGENEOUS OLIGOSACCHARIDE STRUCTURES SEEN IN MATURE PROTEINS Golgi complex - FUNCTIONS GLYCOSYLATION MODIFICATION OF THE N-LINKED OLIGOSACCHARIDES THAT WERE ADDED TO PROTEINS IN THE ER - 3 glucose residues and 1 mannose are removed in the ER - Other sugars are added sequentially by various glycosyltransferases in an ordered sequence of reactions DIFFERENT GLYCOPROTEINS ARE MODIFIED DURING THEIR PASSAGE THROUGH THE GOLGI - Depending on both the structure of the protein and on the amount of processing enzymes that are present within the Golgi complexes of different types of cells  Golgi complex - FUNCTIONS GLYCOSYLATION THE VARIOUS GOLGI CISTERNAE HAS A DISTINCT POPULATION OF RESIDENT ENZYMES Reduced osmium Nucleoside Mannosidase II tetroxide diphosphatase localized in impregnating cis localized in medial cisternae cisternae trans cisternae Golgi complex - FUNCTIONS GLYCOSYLATION  Golgi complex - FUNCTIONS GLYCOSYLATION GLYCOSILATION OF PLASMA MEMBRANE/SECRETED PROTEINS - Removal of 3 additional mannose residues - Sequential addition of an N- acetylglucosamine - Removal of 2 more mannoses - Addition of a fucose and 2 more N- acetylglucosamines - Addition of 3 galactose and 3 sialic acid residues  Golgi complex - FUNCTIONS GLYCOSYLATION GLYCOSILATION OF LYSOSOMAL PROTEINS - Addition of N-acetylglucosamine phosphates to specific mannose residues in the CGN - Removal of the N- acetylglucosamine group, leaving mannose-6-phosphate residues on the N-linked oligosaccharide  Golgi complex - FUNCTIONS GLYCOSYLATION GLYCOSILATION OF LYSOSOMAL PROTEINS - Because of this modification, these residues are not removed during further processing - Instead, these phosphorylated mannose residues are specifically recognized by a mannose-6-phosphate receptor in the TGN, which directs the transport of these proteins to lysosomes Golgi complex - FUNCTIONS Modification and synthesis of the carbohydrate portions of glycoproteins Synthesis of glycolipids and sphingomyelin  Golgi complex - FUNCTIONS LIPID METABOLISM - Sphingomyelin (the only nonglycerol phospholipid in cell membranes) or other glycolipids are synthesized from the conversion of ceramide (synthetized in the ER) Golgi complex - FUNCTIONS Modification and synthesis of the carbohydrate portions of glycoproteins Synthesis of glycolipids and sphingomyelin Processes and sorts proteins received from the RER for further destinations Processes cellular constituents that travel along the secretory pathway