no slide title · comprendono sfingomieline, cerebrosidi e gangliosidi. c) oligosaccaridosi:...

Proteoglycans Are Assembled in the Golgi Apparatus

Modificazioni post-traduzionali delle proteine

La O-glicosilazione

La O-glicosilazione è un processo altamente specifico, che non vede l'aggiunta "seriale" di

carboidrati alla proteina in processazione.

Si svolge completamente nell'apparato del Golgi, dove zuccheri vengono legati al

peptide a livello dell'atomo di ossigeno delle catene laterali di serina o treonina.

L'aggiunta riguarda un singolo carboidrato alla volta e solitamente il numero di zuccheri

legati durante questo processo è limitato a pochi residui.16

Serina o

treonina

Affected gene Protein Affected PTM Disease Major clinical manifestations

MAN1B1 α 1,2 mannosidase glycosylation MAN1B1-CDG severe mental retardation, delayed speech

SLC35A1 CMP-sialic acid transporter glycosylation SLC35A1-CDG seizures, intellectual disability, ataxia, bleeding

SLC35A2 UDP-galactose transporter glycosylation SLC35A2-CDG intellectual disability, seizures, skeletal abnormalities

SLC35A3 UDP-GlcNAc transporter glycosylation Arthrogryposis, mental retardation and seizures autism spectrum disorder, hypotonia, epilepsy, and arthrogryposis

SEC23B Sec23 homolog B glycosylation Dyserythropoietic anemia, congenital, type IIerythroblastic anemia: splenomegaly, gallstones, and iron overload potentially with liver cirrhosis or

cardiac failure.

TRIP11 Golgi microtubule associated protein 210 glycosylation Achondrogenesis type 1A severe chondrodysplasia, lethal before or shortly after birth

UBE3A Ubiquitin ligase E3A glycosylation Angelman syndrome intellectual disability, seizures, lack of speech, and characteristic abnormal behavior

COG2 Component of oligomeric Golgi complex 2 glycosylation COG2-CDGmicrocephaly, developmental delay, intellectual disability, seizures, facial dysmorphism, liver

dysfunction

SLC33A1Solute carrier family 33 (acetyl-CoA

transporter), member 1acetylation Spastic paraplegia-42 spastic gait, increased lower limb tone, weakness and atrophy of the lower limb muscles, pes cavus

CHST3 Chondroitin 6-O-sulfotransferase sulfationSpondylo-epiphyseal dysplasia with joint

dislocationsunusual skeletal dysplasia

CHST6Corneal N-acetylglucosamine-6-O-

sulfotransferasesulfation Macular corneal distrophy type II progressive corneal opacification and reduced corneal sensitivity

CHST8 GalNAc-4-O sulfotransferase I sulfation Peeling skin syndrome general skin peeling

CHST14Dermatan sulfate GalNAc-4-O

sulfotransferase Isulfation

Ehlers-Danlos syndrome musculocontractural

type 1craniofacial dysmorphism, congenital contractures of thumbs and fingers, clubfeet, severe kyphoscoliosis

ARSE Arylsulfatase E sulfation Chondrodysplasia punctata 1 stippled epiphyses, brachytelephalangy, nasomaxillary hypoplasia

PAPPS2 PAPS synthase sulfation Brachyolmia type 4short-trunk stature, rectangular vertebral bodies, precocious calcification of rib cartilages, short femoral

neck. Early death for severe cases.

SLA26A2 Sulfate anion transporter sulfation Achondrogenesis type 1B severe chondrodysplasia, early death of respiratory failure

Atelosteogenesis type 2 pulmonary hypoplasia, lethal in infants

Epiphyseal dysplasia multiple 4 joint pain, scoliosis, malformations of the hands, feet, and knees

Diastrophic dysplasia scoliosis, clubfeet, malformed pinnae with calcification of the cartilage, cleft palate in some cases

GNTPGN-acetylglucosamine-1-

phosphotransferase gamma subunitphosphorylation Mucolipidosis III gamma short stature, skeletal abnormalities, cardiomegaly, and developmental delay

GNTPABN-acetylglucosamine-1-

phosphotransferase alpha and beta subunitsphosphorylation Mucolipidosis II and III

Hip dislocation, gingival hyperplasian, thoracic deformities and hernia soon after birth. Delayed

psychomotor development. Same clinical features for mucolipidosis III as described just above.

IMPAD1 Golgi-resident PAP phosphatase phosphorylation Chondrodysplasia with joint dislocationsshort stature, chondrodysplasia with brachydactyly, congenital joint dislocations, micrognathia, cleft

palate, and facial dysmorphism

INPP5E Inositol polyphosphate-5-phosphatase phosphorylation Morm syndrome Mental retardation, truncal obesity, retinal dystrophy, and micropenis

phosphorylation Joubert syndrome 1Heterogenous: hypoplasia of the cerebellar vermis with the characteristic neuroradiologic molar tooth

sign , dysregulation of breathing pattern and developmental delay.

AKAP9 A-kinase anchor protein 9 phosphorylation Long QT syndrome-11 recurrent syncope, seizure, or sudden death

FAM20CGolgi kinase (family with sequence

similarity 20, member C)phosphorylation Raine syndrome neonatal osteosclerotic bone dysplasia, increased ossification of the skull

CAMKMT Calmodulin-lysine N-methyltransferase methylation 2p21 deletion syndrome cystinuria, neonatal seizures, hypotonia, severe somatic and developmental delay, facial dysmorphism

MBTPS2 Site-2 protease proteolytic cleavageIFAP syndrome with or without BRESHECK

syndromeichthyosis follicularis, atrichia, and photophobia

ZDHHC8 Zinc finger, DHHC-type containing 8 palmitoylation Schizophrenia susceptibilityhallucinations and delusions, inappropriate emotional responses, disordered thinking and concentration,

erratic behavior

ZDHHC9 Zinc finger, DHHC-type containing 9 palmitoylation X-linked mental retardation (Raymond type) general intellectual limitations associated with impairments in adaptive behavior

ZDHHC15 Zinc finger, DHHC-type containing 15 palmitoylation X-linked mental retardation-91 general intellectual limitations associated with impairments in adaptive behavior

PPT1 Palmitoyl-protein thioesterase 1 palmitoylation Neuronal ceroid lipofuscinosis 1Heterogenous: progressive dementia, seizures, and progressive visual deficiency. The cellular phenotype

includes intracellular accumulation of autofluorescent lipopigment storage material.

Degradazione delle macro molecole

EXTRACELLULARI

Funzioni dei lisosomi

(Proteasoma degrada le proteine citosoliche

Funzioni dei lisosomi

(Proteasoma degrada le proteine citosoliche INTRACELLULARI

Degradazione di organelli

Anatomia di un Lisosoma

LisosomiRecupero dei

prodotti

degradati

Mannose 6-phosphate (M6P)

sent to lysosomes

Mannose

How do lysosomal enzymes get to the lysosome?

lysosome

Biosynthesis and traffickingof a lysosomal hydrolase

receptor-dependenttransport

receptor recycling

binding to M6Preceptor

removal of phosphate

addition of phosphate

dissociation at acidic pH

addition of clathrin coat

mature lysosomal hydrolase

Golgi apparatus

Man-6P as a lysosomal targeting signal for proteins with N-linked glycosylation

D-Mannose

Mannose

Cytoplasm ER

Golgi

ASN Transferase absent in I-cell Disease

Receptor is on lysosome bound vesicles but will also end up on the cell surface

Classificate in vari gruppi:

a) Mucopolisaccaridosi: difetto nella degradazione dei mucopolisaccaridi (o GAG),

che svolgono importanti funzioni nel tessuto connettivo.

b) Sfingolipidosi: difetto nella degradazione lisosomiale degli sfingolipidi, che

comprendono sfingomieline, cerebrosidi e

gangliosidi.

c) Oligosaccaridosi: accumulo di oligosaccaridi e di glicoproteine. In

questa classe rientrano la Fucosidosi, la Sialidosi, la Mucolipidosi,

la Mannosidosi.

- Malattie dovute a trasporto lisosomiale alterato.

- Malattie dovute al mancato trasporto degli enzimi lisosomiali.

- Altre malattie lisosomiali (es. Malattia di Pompe, dovuta ad un

deficit di a-glucosidasi).

Malattia

Materiale prevalentemente

accumulato Deficit enzimatico

M. di Gaucher Glucorebroside Glucorebrosidasi

M. di Niemann-Pick Sfingomielina Sfingomielinasi

Leucodistrofia globoide Galattocerebroside Galattocerebrosidasi

Leucodistrofia metacromatica Sulfatide Aril-sulfatasi A

M. Di Fabry Globotriaosilceramide Alfa-galttosidasi A

Fucosidosi Pentaesosilfuco-glicolipide Alfa-fucosidasi

M. Di Farber Ceramide Ceramidasi

Gangliosidosi generalizzata Ganglioside GM1GM1 ganglioside: beta

galattosidasi

M. Di Tay-Sachs Ganglioside GM2 Esosaminidasi A

M. Di SandhoffGanglioside GM2

Esosaminidasi A e B

I-Cell disease (Mucolipidosis Type II)

Autosomal recessive (1/640,000 live birth)

Absence of N-acetylglucosaminyl-1-phosphotransferase.

No Man-6P signal -> enzymes are not targeted to lysosomes -> they are instead secreted to the media

Build up of proteins which cannot be degraded in vacuols. Massive increase in number and size of vacuols which

compromises cell architecture and function.

Overall symptoms

Growth failure and failure to thrive are rapidly progressive.

Developmental delay is severe and often the presenting symptom.

Coarse facial features and musculoskeletal abnormalities

Frequent upper respiratory tract infections

Death from pneumonia or congestive heart failure occurs in early childhood.

I-cell disease patient, 3.5 years old, with her 1week-old sister

Tiny size (not much bigger than her newborn sister)Skeletal abnormalities causing disproportionately large headFingers clenched (joints can’t relax)Gums very swollen

The patient died shortly after this picture was taken.

lysosome

Biosynthesis and traffickingof a lysosomal hydrolase

receptor-dependenttransport

receptor recycling

binding to M6Preceptor

removal of phosphate

addition of phosphate

dissociation at acidic pH

addition of clathrin coat

mature lysosomal hydrolase

Golgi apparatus

Fabry

• Fabry disease is a lysosomal storage disorder seen in one out of every 40,000

• It is caused by a deficiency in the enzyme alpha-galactosidase

• which then results in the body’s inability to break down specific fatty substances called globotriaosylceramide (abbreviated GL-3 or Gb3).

Hurler- Schie

• Mucopolysaccharidosis I is a lysosomal storage disorder that is abbreviated MPS I and sometimes called Hurler syndrome, Hurler-Scheie syndrome, or Scheie syndrome.

• It is caused by a deficiency in the enzyme alpha-iduronidase

• which is needed to break down certain complex sugars called glycosaminoglycans (abbreviated GAGs and formerly called mucopolysaccharides).

MPS IV (sindrome di Morquio)

MPS VI (sindrome di Maroteaux-Lamy)

ENDOCITOSITrasporto vescicolare

Materiale extracellulare è trasportato

all’interno della cellula in vescicole

di endocitosi, rivestite di clatrina,

che gemmano dalla membrana

plasmatica e si fondono con gli

endosomi precoci.

Al livello degli endosomi precoci, i

componenti di membrana che hanno

preso parte al processo , vengono

riciclati e fanno ritorno alla

membrana plasmatica, mentre gli

endosomi precoci si trasformano

gradualmente in endosomi tardivi.

L’endocitosi definisce l’insieme dei processi di

trasporto intracellulare di sostanze o organismi

extracellulari

Tre tipi di endocitosi:

1. Pinocitosi

2. Fagocitosi

3. Endocitosi mediata da recettore

In genere il denominatore comune di questi meccanismi di

internalizzazione di sostanze extracellulari è la formazione

di vescicole più o meno voluminose delimitate da

membrana

Le sostanze raggiungono i

lisosomi attraverso 4 vie

• Pinocitosi generalizzata

– Cellula “beve” < 150 nm

• Endocitosi Mediata da Recettore

• Fagocitosi

– Cellula “mangia” > 250 nm

• Autofagia

– Cellula “mangia se stessa” organelli invecchiati o non

più funzionali

– Meccanismo importante nello sviluppo, nell’omeostasi

e nella morte cellulare

Sostanze disciolte in acqua (soluti) sono continuamente

internalizzate dalle cellule via Pinocitosi.

Piccole invaginazioni della membrana plasmatica formano

microvescicole conteneti i soluti che progressivamente

vengono rilasciati nel citoplasma

Endocitosi

• Pinocitosi

– Letteralmente: la cellula beve

– Importo di fluidi che vengono

rilasciati nel citosol

Pinocytosis(a form of endocytosis)

2. FAGOCITOSI

FAGOCITOSI

Escape of Listeria monocytogenes from a Vacuole

Autophagy Degrades Unwanted Proteins and Organelles

Autophagy Degrades Unwanted Proteins and Organelles

Autophagy Degrades Unwanted Proteins and Organelles

clathrin coated

pitvesicle

IN

OUT

IN

OUT

IN= cytosol

3. receptor mediated endocytosis

adaptin

cargo receptor

OUT

IN

clathrin

OUT

IN

+

dynamin

OUT

IN

clathrin coated vesicle

OUT

IN

GTP GDP + Pi

pinching off

uncoating

ATP ADP + Pi

clathrin coated vesicle

OUT

IN

cytosol

v-SNAREt-SNARE

docking

OUT

IN

Endosome

fusion

Receptor-mediated endocytosisEndosome

cytosol

OUT

clathrin triskelions clathrin cage

Michael Brown

Joseph Goldstein

Noble Prize 1985

"for their discoveries concerning the

regulation of cholesterol metabolism"

LDL

= low density lipoprotein

cholesterol

out

cytosol

Receptor recyclingout

pH: 5.5

out

cytosol

Hypercholesterolemia

Atherosclerosis

The mutations

• Five classes

– I – can’t synthesize LDLR

– II – Targeting mutation

– III – can’t bind LDL

– IV – can’t cluster receptor in clathrin coated pit and internalize

– V – can’t recycle LDL

EarlyEndosome

Transportvesicle

Recyclingvesicle

Microtubule

Endosomecarrier vesicle Dynein

Degradation of the Ligand and Recycling of its Receptor

COP-I

LDL LDL receptor

Actin

Recyclingvesicle

EarlyEndosome

Transportvesicle

Microtubule

Endosomecarrier vesicle Dynein

Fe

Recycling of the Ligand and its Receptor

Transferrin-Fe Transferrin

Actin

Receptor

EarlyEndosome

Transportvesicle

Recyclingvesicle

Microtubule

Endosomecarrier vesicle Dynein

Transcytosis of the Ligand and its Receptor

IgG

Many Proteins and Lipids Are Carried Automatically from the Trans Golgi Network (TGN) to

the Cell Surface

Secretory Vesicles Bud from the Trans Golgi Network

secretion

retrovirus

15.9

Viral env