CSB327 Lecture 1 Notes

CSB327 Lecture 1 Notes – Introduction (September 10, 2012) 1 – Extracellular matrix dynamics and associated pathologies • Jellyfish form a special matrix between two cell layers that allow them to move freely and have strength. Spider webs have strength and elasticity. One of the most abundant group of molecules in our body are collagens. They provide tensile strength. It wasn’t that long ago that people perceived ECM as primarily structural entities, to which cells bound to as a scaffold. • An ECM is an integrated circuit of complex matrix molecules. 9 – Matrisome approximately • If you look at the sum total of genes that code for matrix molecules, there are over 300 genes in your bodies that code for different types of ECM molecules. All of these molecules have to be made in the right way, assembled into a proper scaffold, and communicating with the self. It is a complex environment of molecules. 10 – Two major types of ECM • There are two main types of ECM in our bodies. The basal lamina is a special extracellular sheet that plays a barrier function and an adhesive function. In most cases, the basal lamina is called the basement membrane. The basal lamina is a specialized sheet that underlies epithelia, surrounds muscle cells, adipose cells and peripheral nerves. The basal lamina is universal in the animal kingdom. There isn’t always as much as the connective tissue. The connective tissue ECM or interstitial ECM is a thick layer that underlies the basal lamina. We are going to look at the molecules that make up both of these. Collagen fibers are found in basal lamina. The connective tissue is a composite of all the major types of ECM molecules that we find in the body. Depending on the type of connective tissue, you will find different distributions of the ECM molecules, different isoforms that give the tissues their specific shape, biomechanical properties, physiological function and cellular activities. • There are two major types of matrices: basal lamina/basement membrane and interstitial matrices. Collagens are found in both matrices. 11 – Basal lamina in the cornea of a chick embryo • The basal lamina is a sheet like network. You can see the collagen fibrils underlying the basal lamina. The majority of cancers are derived from epithelial cells. The basal lamina is your best friend. Until melanomas traverse the basal lamina and enter the vascular network, then you are fine. Cancer cells can break and create holes in the basal lamina to which they are then free to travel. If you catch the cancer early enough and you have the protection of the basal lamina, then you are fine. The early diagnosis of melanomas is critical. The cells are in an intimate contact with the basal lamina. This is a major adhesive sheet that promotes anchoring of cells and migration along the basal lamina. The basal lamina is an intimate contact with the underlying connective tissue, collagen network and other ECM molecules. The basal lamina stabilizes the interface between the epidermis and the dermis. Mutations in the genes that affect the basal lamina will separate the epidermis from the dermis. There are two major classes of ECM: basal lamina and interstitial ECM. Both are enriched in different types of collagens. 12-14 – Lecture topics • This is a tentative list of lectures that I will follow. 15 – Classes of secretory proteins in vertebrates • Do not memorize this slide. • What is an ECM molecule? There are a lot of molecules outside your cells that we don’t classify as ECM. These ECM molecules are made by all tissues. We used to think our brain did not make any ECM molecule. During early neurogenesis, cells are moving rapidly. ECM molecules can create a permissive environment to guide the axons. There are some non-collagenous structural proteins like lamprin and egg shell proteins that are similar in molecular composition to elastin but they do not have elastic properties. 17 – In addition to forming elaborate 3D scaffolds with complex biomechanical properties, ECM molecules have diverse regulatory functions that affect cell • If we look at elaborate 3D scaffolds with complex biochemical properties, ECM molecules have diverse regulatory functions that affect all cellular behaviour. We will find that cells embedded inside interstitial matrices will commit suicide by apoptosis if you take them out of their matrices. Their survival depends on ECM molecules. We will see that you not just have attachment, but regulatory function too. 18 – Dynamic reciprocity or cross-talk • ECM molecules are critical to the development of multicellular organisms. We will see that vertebrate and invertebrate embryos begin to make a matrix very quickly following cellular division. Dynamic reciprocity is communication cross-talk between the matrix being made. The bone is a specialized connective tissue. The bone has to adapt to changes in development. What the cell makes in a matrix will change the biological activity of the cell that made that matrix, which in turn will change the matrix that it made and remodel that so that you have continuous growth that is synchronized with development. Matrix homeostasis is fundamental to development and this involves continuous remodelling during early development. 19 • I don’t know of any animal that doesn’t make matrix. Sponges do not have a nice matrix. Jellyfish make an excellent matrix. The evolution of animals is dependent on the generation of new genes that constitute the matrix. 20 – Examples of pathologies associated with mutations in ECM molecules • The cartilage is the first skeleton that you make which sets up the future shape of the bone. If you mess up the cartilage, then you get achondroplatic dwarfism. If you have defects in the collagen, then you get osteogenesis imperfect. If the matrix components that make up the joints are shared with the vasculature, then you get EDS. 21 – How are ECM molecules secreted? • How are ECM molecules secreted without any interference to the outside of the cell? If you started making matrix inside your cells with molecules that are capable of mineralizing tissue, then your cells would solidify. You have a lot of calcium phosphate in ECM. It is critical that all the events that occur in the ER go right, otherwise the end product is compromised. This is an epistatic event. • There is no single ECM molecule that does not impact on the biological activity of cells and how they behave. Dynamic reciprocity is continuous cross-talk between the matrix and the cells that make them. You need to make a matrix that will give some shape. During evolution, a new set of genes were born from ancestral forms. As evolution progressed, new genes were made and modified so that you have specialized function and adaptations to the environment. Without the ECM would be an enormous mass of cells. You wouldn’t have any shape or biomechanical properties. ECM was perceived as boring material and static. • I am using pathologies to emphasize the importance of individual assemblies. I will start with type I collagen because type I collagen is the prototype or most abundant in our body. How are ECM molecules secreted? What are the consequences of failure to move from the cytoplasm to the matrix? 22 – Overview of intracellular protein traffic • These molecules need a synergistic interaction with the matrix that surrounds the cells. This is the trafficking of proteins inside a cell. We are going to the ER. How does a cell make a decision to ship a protein to the ER? There is a special protein sequence called a signal peptide that will play a critical role. The secretory pathway is where I am focused on. I will not ask you anything about trafficking in the other ways. What precedes mRNA? What do you make first? What is the first transcript that is made called? Heterogeneous nuclear RNA (hnRNA) gets spliced and capped. If you are going to make a molecule, especially collagen, which has the potential to form fibrils, then you do not want that occurring inside the cell. mRNA that are fated to go to the ER will undergo translation arrest before they start translation. Before the ER, they will arrest translation. I will focus on translation arrest and delivery to the ER today. 23 – Anatomy of cell • This is a schematic of the cell to emphasize all the different compartments. 24 – Synthesis of a protein destined for the ER • Translation begins in the cytosol. The ribosome has a translation start site and there is a start codon to begin translation. When you have made about 70 amino acids, a portion of that nascent peptide will be sticking out of the ribosome. This is very critical. This is when the sequence called a signal peptide becomes exposed outside of the ribosome. As a consequence, the signal peptide is recognized by a signal receptor particle. Often in literature, a particle refers to a composite of nucleic acids and protein. A signal receptor particle will see the signal peptide and bind to it. What is special about the signal peptide? What are the universal features to recognize a signal peptide anywhere along the secretory pathway? 25 – Amino acid sequences of signal peptides of several eukaryoti