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Flashcards in Proteoglycans and Integrins Deck (20):
1

Proteoglycans in the ECM The study of proteoglycans dates back to the beginning of the 20th century with investigations of “chondromucoid” from cartilage and anticoagulant preparations from liver (heparin)

The proteoglycan molecules in connective tissue form a highly____, gel-like “___ ____” in which the fibrous ____are embedded

The polysaccharide gel resists ____ forces on the matrix while permitting the rapid ____ of nutrients, metabolites, and hormones between the___ and the tissue ___.

Proteoglycans consist of _____ (called _____) linked to a core ____

Note – Non-ECM proteoglycans are _______and can either ___ the plasma membrane (type I membrane proteins) or are linked by a ___ ___.

The study of proteoglycans dates back to the beginning of the 20th century with investigations of “chondromucoid” from cartilage and anticoagulant preparations from liver (heparin) The proteoglycan molecules in connective tissue form a highly hydrated, gel-like “ground substance” in which the fibrous proteins are embedded The polysaccharide gel resists compressive forces on the matrix while permitting the rapid diffusion of nutrients, metabolites, and hormones between the blood and the tissue cells Proteoglycans consist of polysaccharides called glycosaminoglycans (GAGs) linked to a core protein Note – Non-ECM proteoglycans are membrane-bound and can either span the plasma membrane (type I membrane proteins) or are linked by a lipid anchor.

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Glycosaminoglycans

GAGs are _____polysaccharide chains composed of ____ disaccharide units

One of the two sugars in the repeating disaccharide is always an ___ sugar (____ or ____), which in most cases is ____.

 The second sugar is usually a ____ ____ (____ or ____) or ____

Because there are___ or ____ groups on most of their sugars, GAGs are highly ____ charged (the most anionic molecules produced by animal cells)

Because of the negative charges, proteoglycans bind ____ charged ions and form _____ _____ with trapped water molecules, thereby creating a hydrated gel S

even main groups of GAGs are distinguished according to their ____, the type of ____ between the sugars, and the___ and ____ of ____ groups:

(1) ____, (2) ___ ___, (3) ___ ___, (4) ___, (5) ___ ____, (6) ___ __ __and (7) ___ ___ __

_____ is the only GAG that is not sulfated

GlycosaminoglycansGAGs are unbranched polysaccharide chains composed of repeating disaccharide units One of the two sugars in the repeating disaccharide is always an amino sugar (N-acetylglucosamine or N-acetylgalactosamine), which in most cases is sulfated The second sugar is usually a uronic acid (glucuronic or iduronic) or galactose Because there are sulfate or carboxyl groups on most of their sugars, GAGs are highly negatively charged (the most anionic molecules produced by animal cells) Because of the negative charges, proteoglycans bind positively charged ions and form H-bonds with trapped water molecules, thereby creating a hydrated gel Seven main groups of GAGs are distinguished according to their sugars, the type of linkage between the sugars, and the number and location of sulfate groups: (1) hyaluronan, (2) chondroitin sulfate, (3) dermatan sulfate, (4) heparin, (5) heparan sulfate, (6) Keratan sulfate I and (7) keratan sulfate II Hyaluronan is the only GAG that is not sulfated

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Synthesis and structure of proteoglycans

Structure of proteoglycans resemble a ___ ___ with many GAG chains extending from a core proteins

The core protein is made on ___ ___ ribosomes and threaded into the____of the ER

Glycosylation occurs initially as the protein enters the ___ ____ and then in the ___

____ sugars act as precursors and add ___ ___to the protein and then to the growing end of the ____ chain

The majority of GAGs linked to protein to form a proteoglycan are attached via a _____ linker that consist of ___ ___ ___ ___ - ___ residue in the protein core

Hyaluronan does not occur _____ linked to proteoglycans, but instead interacts ____ with proteoglycans via hyaluronan-___ ____; other GAGs attach to the core protein ____.

Except ___ ____, all GAGs are attached to proteins via a ___ or____residue; keratan sulfate is attached to ____.

 Two types of _______ are the enzymes responsible for addition of ___.

Sulfation occurs after ____ of ___.

After synthesis, proteoglycans are ____ from the cells

Structure of proteoglycans resemble a bottle brush with many GAG chains extending from a core proteins The core protein is made on membrane-bound ribosomes and threaded into the lumen of the ER Glycosylation occurs initially as the protein enters the ER lumen and then in the golgi UDP sugars act as precursors and add sugar units to the protein and then to the growing end of the carbohydrate chain The majority of GAGs linked to protein to form a proteoglycan are attached via a tetrasaccharide linker that consist of GlcA–Gal–Gal–Xyl–Ser residue in the protein core Hyaluronan does not occur covalently linked to proteoglycans, but instead interacts noncovalently with proteoglycans via hyaluronan-binding motifs; other GAGs attach to the core protein covalently. Except keratan sulfate, all GAGs are attached to proteins via a serine or threonine residue; keratan sulfate is attached to asparagine Two types of glycosyltransferases are the enzymes responsible for addition of sugars Sulfation occurs after addition of sugars After synthesis, proteoglycans are secreted from the cells

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Proteoglycans vary in size

Proteoglycans can contain as much as ___% carbohydrate by weight, mostly in the form of long, ____ GAG chains, each typically about 80 sugars long

The proteoglycan aggrecan, which is a major component of cartilage, has a mass of about 3 × 106 daltons with over ____ GAG chains

Few proteoglycans are much smaller and have only ____ GAG chains; decorin, which is secreted by fibroblasts, has a__GAG chain.

Proteoglycans can contain as much as 95% carbohydrate by weight, mostly in the form of long, unbranched GAG chains, each typically about 80 sugars long The proteoglycan aggrecan, which is a major component of cartilage, has a mass of about 3 × 106 daltons with over 100 GAG chains Few proteoglycans are much smaller and have only 1–10 GAG chains; decorin, which is secreted by fibroblasts, has a single GAG chain.

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Aggrecan

A number of _____ interact with ____ to form large complexes

A well-characterized example is aggrecan

Core protein of aggrecan (shown in red line) contains:

  • ____-terminal domain that can bind ____  with the help of (____protein)
  • a central domain to which ____ sulfates and ____ sulfates are attached
  • a carboxy-terminal domain that contains a _-____ _____ domain

A number of proteoglycans interact with hyaluronan to form large complexes A well-characterized example is aggrecan Core protein of aggrecan (shown in red line) contains: amino-terminal domain that can bind hyaluronans with the help of (link protein) a central domain to which chondroitin sulfates and dermatan sulfates are attached a carboxy-terminal domain that contains a C-type lectin domain

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General functions of ECM proteoglycans

Provide ____ space around and between cells

Their GAG chains can form gels of varying ___size and charge density and thereby serve as selective sieves to regulate the ___ of ____ and cells according to their ___ , ____, or both

Through interaction with collagen, proteoglycans aid in the ___ ____ of most tissues

Bind various ___ ___ molecules, such as certain protein __ ___, and enhance (by preventing their ____) or___their signaling activity

Proteoglycans bind and regulate the activities of, other types of secreted proteins, including ___ ____ (proteases) and ___ ____

Provide hydrated space around and between cells Their GAG chains can form gels of varying pore size and charge density and thereby serve as selective sieves to regulate the traffic of molecules and cells according to their size, charge, or both Through interaction with collagen, proteoglycans aid in the structural organization of most tissues Bind various secreted signal molecules, such as certain protein growth factors, and enhance (by preventing their proteolysis) or inhibit their signaling activity Proteoglycans bind and regulate the activities of, other types of secreted proteins, including proteolytic enzymes (proteases) and protease inhibitors

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Specific functions proteoglycans

___: Cell ___

___: Formation of___ ___ ___

__: ___of cornea

__:____of cornea, Binds ___

____: Anticoagulant, cause release of ___ ____

___: Component of skin fibroblast and aortic wall

Specific functions proteoglycans

HA: Cell migration

CS: Formation of bone cartilage, cornea

KS: Transparency of cornea

DS:Transparency of cornea, Binds LDL

Heparin: Anticoagulant, cause release of Lipoprotein Lipase

HS: Component of skin fibroblass and aortic wall

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Degradation of proteoglycans

ECM proteoglycans are ____

Endocytic vesicles fuse with ____

Lysosomal ___and____ degrade the core protein and carbohydrate components of the proteoglycans, respectively

_______ are a group of metabolic disorders caused by the ___ or ____ of different lysosomal enzymes needed to break down GAGs – results in ____ deformities

Degradation of proteoglycansECM proteoglycans are endocytosed Endocytic vesicles fuse with lysosomes Lysosomal proteases and glycosidases degrade the core protein and carbohydrate components of the proteoglycans, respectively Mucopolysaccharidosis are a group of metabolic disorders caused by the absence or malfunctioning of different lysosomal enzymes needed to break down GAGs – results in skeletal deformities

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Interns

The linkage of the extracellular matrix to the cell requires transmembrane ___ ___ ___ that act as matrix ____ and tie the ___ to the cell's ___

The principal receptors on animal cells for binding most extracellular matrix proteins—including ___ ____ ____—are the ___

Integrins play central roles in the biology by controlling___ ____ to the ECM and cell _____, ___, ____ and ____.

As a result, they contribute to the regulation of ____, ___, ____ and ___, and to the development of ___

 Integrins differ from other cell surface receptors because they:

Bind their ligands with much____ affinities Are expressed ____fold more Why? If the binding were too tight, cells would become ___ ___ to the matrix and would be unable to ____—a problem that does not arise if attachment depends on large numbers of weak adhesions -“___ principle”

The linkage of the extracellular matrix to the cell requires transmembrane cell adhesion proteins that act as matrix receptors and tie the matrix to the cell's cytoskeleton The principal receptors on animal cells for binding most extracellular matrix proteins—including collagens, fibronectin and laminins—are the integrins Integrins play central roles in the biology by controlling cell adhesion to the ECM and cell migration, growth, differentiation and apoptosis.

As a result, they contribute to the regulation of development, immunity, inflammation and hemostasis, and to the development of diseases Integrins differ from other cell surface receptors because they: Bind their ligands with much lower affinities Are expressed 10-100 fold more Why? If the binding were too tight, cells would become irreversibly glued to the matrix and would be unable to move—a problem that does not arise if attachment depends on large numbers of weak adhesions -“Velcro principle”

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Structure of integrins

An integrin molecule is composed of ___ ______associated transmembrane ___ subunits called __ and __ 

The heterodimer integrin consists of a ____ extracellular (___) ligand-binding head, ___ ____-pass transmembrane helices and ___ ___cytoplasmic tails

A ____ of human integrin heterodimers are formed from 9 types of β subunits and 24 types of α subunits

Different integrins can bind the___ ____ ____. Ex: At least 5 different integrins bind laminin

xAn integrin molecule is composed of two noncovalently associated transmembrane glycoprotein subunits called α and β The heterodimer integrin consists of a large extracellular (ecto) ligand-binding head, two single-pass transmembrane helices and two short cytoplasmic tails A variety of human integrin heterodimers are formed from 9 types of β subunits and 24 types of α subunits Different integrins can bind the same ECM protein. Ex: At least 5 different integrins bind laminin

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Activation of integrins and signaling

Integrins exist in an ____ between a __ ____-affinity state and an ___ ___-affinity state

When integrins are in the resting state, the α and β ____ tails are in ____ proximity, which constrains the integrin in a ____ _____conformation, in a bent form in which the head group points ____towards the cell surface

____ signals can activate the integrins and force them to “___ ___” (___-___ signaling)

During 'inside–out' signaling, an intracellular activator binds to the __ ___ ___, leading to conformational changes that result in ____affinity for extracellular ligands

In 'outside–in' signaling, binding of integrins to their___ ___ changes the conformation of the integrin and, because many of the ligands are multivalent, contributes to ___ ___.

Therefore, integrins are _____ signaling proteins. Inside-out and outside-in signaling occur in consortium

Activation of integrins and signaling Integrins exist in an equilibrium between a bent low-affinity state and an upright high-affinity state When integrins are in the resting state, the α and β cytoplasmic tails are in close proximity, which constrains the integrin in a low-affinity conformation, in a bent form in which the head group points inwards towards the cell surface Intracellular signals can activate the integrins and force them to “stand up” (inside-out signaling) During 'inside–out' signaling, an intracellular activator binds to the β-integrin tail, leading to conformational changes that result in increased affinity for extracellular ligands In 'outside–in' signaling, binding of integrins to their extracellular ligands changes the conformation of the integrin and, because many of the ligands are multivalent, contributes to integrin clustering Therefore, integrins are bidirectional signaling proteins. Inside-out and outside-in signaling occur in consortium

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Functions controlled by integrin signaling

If bound to ECM components: _____ _____

If bound to ligand: ____ _____

If no ligand bound: ____

Functions controlled by integrin signaling

If bound to ECM components: Cell Adhesion Cell Migration
Cell Proliferation, survival and death

If no ligand boundà Apoptosis

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Integrin-mediated cell adhesion and migration

Attachment of cells to ECM components induces ____ of integrins on the cell surface.

The cytoplasmic portions of the clustered integrins then act as a platform for the _____ of ____ proteins such as adaptor/scaffold and signaling proteins to the inner surface of the plasma membrane, where they form structures called ____ ____.

The adaptor/scaffold proteins in FAs provide ____ to the ___ ___ and, thereby, connect cells firmly to the ECM.

This linkage enables the generation of the ____ necessary to alter cell ____ and the____ force necessary to ___ the cell body during migration.

In addition, multiple signaling proteins, including __ or ____, are also recruited to FAs where they transmit ___-derived signals to cellular pathways controlling ___ ____ ____

Attachment of cells to ECM components induces clustering of integrins on the cell surface. The cytoplasmic portions of the clustered integrins then act as a platform for the recruitment of cellular proteins such as adaptor/scaffold and signaling proteins to the inner surface of the plasma membrane, where they form structures called focal adhesions. The adaptor/scaffold proteins in FAs provide linkages to the actin cytoskeleton and, thereby, connect cells firmly to the ECM. This linkage enables the generation of the tension necessary to alter cell morphology and the traction force necessary to move the cell body during migration. In addition, multiple signaling proteins, including kinases or phosphatases, are also recruited to FAs where they transmit ECM-derived signals to cellular pathways controlling proliferation, survival and migration.

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Visualizing focal adhesion formation and actin dynamics in adhering and migrating cells

Visualizing focal adhesion formation and actin dynamics in adhering and migrating cells

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Armed to the teeth with αvβ6 intern

Tooth enamel is the hardest mineralized tissue in the body and is produced by cells called ____.

During the secretory stage of amelogenesis, enamel proteins such as ____ are secreted into the enamel ____, which is in direct contact with the____ plasma membrane.

A recent study shows that integrin αvβ6 is expressed in _____ where it regulates ____ of ____ and subsequent enamel _____.

In mice deficient in αvβ6 integrin (Itgb6−/−), the researchers found ___ ____ ___,with a significantly reduced ___:____ratio, whereas the molars of these mice show reduced ____ and severe___.

Tooth enamel is the hardest mineralized tissue in the body and is produced by cells called ameloblasts. During the secretory stage of amelogenesis, enamel proteins such as amelogenin are secreted into the enamel matrix, which is in direct contact with the ameloblast plasma membrane. A recent study shows that integrin αvβ6 is expressed in ameloblasts where it regulates deposition of amelogenin and subsequent enamel biomineralization. In mice deficient in αvβ6 integrin (Itgb6−/−), the researchers found chalky rounded incisors, with a significantly reduced mineral-to-protein ratio, whereas the molars of these mice show reduced mineralization and severe attrition.

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Teeth from 6-month-old Itgb6−/− mice show severe attrition.

Teeth from 6-month-old Itgb6−/− mice show severe attrition.

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Integrins and diseases of the oral cavity

Integrins in periodontal disease

Integrin αvβ6 is expressed in the ___ ____ of the gingiva. Animals deficient in this receptor or unable to activate this receptor develop classical signs of ___ ___ suggesting that it plays a role in the regulation of ___ ____.

Patients with mutations in ___-_, a protein that activates these integrins in ___-___ signaling, have severe ___-___ periodontal disease.

Integrins in periodontal disease Integrin αvβ6 is expressed in the junctional epithelium (JE) of the gingiva. Animals deficient in this receptor or unable to activate this receptor develop classical signs of periodontal disease suggesting that it plays a role in the regulation of periodontal inflammation. Patients with mutations in kindlin-1, a protein that activates these integrins in inside-out signaling, have severe early-onset periodontal disease.

18

Integrins and diseases of the oral cavity

Integrins in oral squamous cell carcinoma (OSCC)

Integrin αvβ6 is ___ ____ in OSCC

Overexpression of αvβ6 correlates with increased ____ and ____ of the OSCC cells.

Integrins in oral squamous cell carcinoma (OSCC) Integrin αvβ6 is over expressed in OSCC Overexpression of αvβ6 correlates with increased proliferation and invasiveness of the OSCC cells.

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Integrins and cancer

Integrins expressed in tumor cells contribute to tumor ____ and ____ by increasing tumor cell ____, ____, ____ and ____.

Integrin adhesion to the extracellular matrix (ECM) provides the traction required for tumor cell ___.

 Integrins also contribute to tumor cell invasion by regulating the localization and activity of matrix-_____ proteases, such as ________.

Integrins also regulate proliferation. Integrin ligation controls the expression of key ___ ___ ___, including cyclin __ and the cyclin-dependent kinase _____ family, which regulate entry into the ___ phase of the cell cycle.

Integrins also control tumor cell survival. Ligated integrins prevent ______signaling cascades or _____-mediated death and increase survival signaling.

Integrins expressed in tumor cells contribute to tumor progression and metastasis by increasing tumor cell migration, invasion, proliferation and survival. Integrin adhesion to the extracellular matrix (ECM) provides the traction required for tumor cell invasion. Integrins also contribute to tumor cell invasion by regulating the localization and activity of matrix-degrading proteases, such as matrix metalloprotease 2 (MMP2). Integrins also regulate proliferation. Integrin ligation controls the expression of key cell cycle proteins, including cyclin D1 and the cyclin-dependent kinase inhibitor family, which regulate entry into the S phase of the cell cycle. Integrins also control tumor cell survival. Ligated integrins prevent pro-apoptotic signaling cascades or integrin-mediated death and increase survival signaling.

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Integrin–growth factor and integrin–cytokine receptor crosstalk in cancer

Cooperation between ___ and____ ____ signaling or ___ and ____ signaling is crucial to tumor progression.

Integrin ligation may lead directly to the increased secretion of ___ ___ and/or ____, which can then bind to their receptors in an ___ or ____ manner to further ____ signaling.

Signaling induced by either ___ ___ or __ ___binding may activate common downstream pathways resulting in ___ ____overall compared with the activation of either receptor alone.

Alternatively, both ___ and ___ ___signaling may regulate integrin function by directly controlling integrin ____ levels.

Cooperation between integrin and growth factor signaling or integrin and cytokine signaling is crucial to tumor progression. Integrin ligation may lead directly to the increased secretion of growth factors and/or cytokines, which can then bind to their receptors in an autocrine or paracrine manner to further induce signaling. Signaling induced by either integrin ligation or growth factor binding may activate common downstream pathways resulting in enhanced signaling overall compared with the activation of either receptor alone. Alternatively, both chemokine and growth factor signaling may regulate integrin function by directly controlling integrin expression levels.