The endoplasmic reticulum through proteomics- Identifying the links between morphology and function

The eukaryotic cell is organized into several membranous subcellular compartments that contribute to the spatial segregation of the many cell physiological functions. One of these organelles, the endoplasmic reticulum (ER), is a continuous compartment, emanating from the nuclear envelope, through the rough ER, and ending with the reticulated smooth ER network. Each domain of the organelle is distinct for its morphology, as well as the many diverse functions that it performs. In this thesis, a survey of a proteomics resource for both the rough and smooth domains of the ER reveals that the organelle is further divided into spatial and functional subdomains. Furthermore, the assignment of novel proteins that were uncovered by proteomics to these various functional clusters can provide insight into their functions and guide future functional studies. Lastly, this thesis demonstrates the importance of the distinct morphology of the smooth ER network in maintaining basic physiological processes. Taken together, these data demonstrate that while the ER is a single, continuous, subcellular compartment, it is highly complex in its spatial, functional, and morphological organization.La cellule eucaryote est constituée de plusieurs structures spécialisées dénommées organites qui contribuent à la démarcation des fonctions physiologiques de la cellule. L'un de ces organites, le réticulum endoplasmique (RE), est une structure continue qui provient de l'enveloppe nucléaire, devient le RE rugueux, et se termine avec le réseau réticulé RE lisse. Chaque division de l'organite est unique pour sa morphologie, ainsi que ses fonctions nombreuses et diverses. Dans cette thèse, une analyse d'une ressource protéomique révèle que les deux domaines du RE sont encore divisés en de nouvelles sections régionales et fonctionnelles. De plus, l'attribution de nouvelles protéines découvertes par la protéomique à ces divisions fonctionnelles peut donner un aperçu de leurs fonctions et guider de nouvelles études. Enfin, cette thèse démontre l'importance de la morphologie distincte du réseau RE lisse pour le maintient de certains processus physiologiques de base. Le tout démontre que même si le RE est en effet un compartiment continu, son organisation régionale, fonctionnelle et morphologique est très complexe

Compartmentalization of membrane trafficking, glucose transport, glycolysis, actin, tubulin and the proteasome in the cytoplasmic droplet/Hermes body of epididymal sperm

Discovered in 1909 by Retzius and described mainly by morphology, the cytoplasmic droplet of sperm (renamed here the Hermes body) is conserved among all mammalian species but largely undefined at the molecular level. Tandem mass spectrometry of the isolated Hermes body from rat epididymal sperm characterized 1511 proteins, 43 of which were localized to the structure in situ by light microscopy and two by quantitative electron microscopy localization. Glucose transporter 3 (GLUT-3) glycolytic enzymes, selected membrane traffic and cytoskeletal proteins were highly abundant and concentrated in the Hermes body. By electron microscope gold antibody labelling, the Golgi trafficking protein TMED7/p27 localized to unstacked flattened cisternae of the Hermes body, as did GLUT-3, the most abundant protein. Its biogenesis was deduced through the mapping of protein expression for all 43 proteins during male germ cell differentiation in the testis. It is at the terminal step 19 of spermiogenesis that the 43 characteristic proteins accumulated in the nascent Hermes body