Lecture 2: Nucleus

• MCB 2050


  Course code
• Lecture 1 Review
• Course information - Lecture materials on CourseLink, Discussions page, and recommendations for success (e.g., keep up with lectures, review, tutorials, and SLGs)
• Lecture schedule – Regulation of Gene Expression: Protein synthesis, trafficking & organelle biogenesis
• Nucleus/Microscopy/Endomembrane System/Mitochondria & Chloroplasts

  
• Visualization of spatial organization and timing of gene expression in a living cell

  
• Key molecules


  • DNA
  • Protein
  • mRNA
• Cell Biology Today -"The Post-Genomics, Cellular Imaging Revolution"
• Advances in


  • Functional genomics
  • other '-omics'
  • Stem cells, regeneration biology, artificial cells, cellular basis of disease/applied therapeutics
  • Autofluorescent proteins
  • Imaging/microscopy
• Nucleus – structure and function

  
• Nuclear 'compartmentation'


  • nucleoplasm (nuclear subdomains), nuclear matrix, nuclear envelope, and nuclear lamina
• Nuclear pore complex - structure
• Suggested E-Textbook Readings: Morris Chapter 5 pg 104-115 (Review of 'Cellular Organization'), Lodish Chapter 13 pg 614-620
• Midterm Exam is Saturday - see 'Announcements' on CourseLink for details
• SLG sessions available this week for preparing for midterm
• Tutorial #5 'Nucleus and Nuclear Transport'
• Tutorial readings & application assignment posted next Friday, Mar 8th
• 'Key Words: Nucleus and Nuclear Transport' - available on CourseLink
• Nucleus


  Largest organelle – typically one nucleus per cell
• Nuclear size


  • Varies from cell-to-cell and between organisms, usually determined by cell size (i.e., cytoplasmic volume), increases during development and in cancer cells – used for cancer diagnosis/prognosis
• Primary difference between prokaryotes and eukaryotes

  Eukaryotes possess a membrane-bound nucleus

  
• Eukaryotic nucleus


  • Nuclear envelope – boundary between cytoplasm and nucleus, Nuclear pores – 'doorways' in nuclear envelope; regulate transport in/out of nucleus, Nucleolus - site of ribosome synthesis, Nuclear matrix – fibrillar protein 'mesh' (network); serves in structural support & chromatin scaffold, Nucleoplasm - ordered architecture; site where chromatin is found and RNA processing

  
• Prokaryotic 'region'


  Nucleoid where chromosome is located; less DNA, less DNA packaging, and limited/no RNA processing

  
• Differences between prokaryotes and eukaryotes

  • Eukaryotes: larger, structurally & functionally more complex interiors, possess single and double membrane-bound organelles, Cellular "compartmentalization" allows for larger size and segregation and organization of specific cellular functions, Each organelle contains both unique and common factors for functioning (e.g., metabolism) and their biogenesis (formation), maintenance, and turnover

  
• See Morris textbook (Chapter 5: pg 104-115) for review of general animal and plant cellular organization
• Nucleus - 2 main functions

  1. Compartmentalization of the cellular genome and its activities (e.g., site of DNA replication, transcription & RNA processing, site where translation components (ribosomes, mRNAs, tRNAs) are synthesized), 2. Coordination of cellular activities (e.g., control of metabolism, protein synthesis, reproduction (cell division), etc.)
• Separation of the cytoplasm from the genome allows for unique spatial & temporal regulation of gene expression in eukaryotes

  
• Nucleus (nuclear envelope) limits access of transcription factors (proteins) from cytoplasm to genome
• Nucleoplasm


  Fluid-filled interior of nucleus – highly organized, consists of >30 specialized regions ('subdomains') that participate in specific functions (Note: nuclear subdomains are not membrane bound)

  
• Nucleolus


  Most conspicuous nuclear subdomain - irregular shaped, dense and granular in appearance (via EM), size and number (1-5 nucleoli) depend on metabolic activity of cell (↑cellular activity ↑protein synthesis ↑size/number of nucleoli), function in producing ribosomes (site of ribosomal DNA (rDNA) gene transcription, rRNA processing, and initial stages of ribosomal subunit (rRNA + protein) assembly, final assembly of ribosomes (used for protein synthesis) occurs in cytoplasm

  
• Location of a gene is often related to its activity - most actively transcribed genes (i.e., decondensed euchromatin) are found at periphery of chromosomal 'subdomains'
• Interchromosomal channels – regions between chromosome subdomains that serve as barriers to prevent unwanted DNA-DNA and/or DNA-protein interactions
• Active genes (euchromatin) from different subdomains (or from different regions of same chromosome) extend into interchromosomal channels to form transcription factories where transcription factors are concentrated
• Interchromosomal interactions – "kissing chromosomes", gene regulatory regions from one chromosome activate gene(s) on another chromosome
• Nuclear speckles


  Subdomains (appear as 'speckles' via fluorescence microscopy) where mRNA splicing factors concentrated (i.e., where pre-mRNA processing occurs), often located in interchromosomal channels next to transcription factories, numerous and highly dynamic – often move quickly and grow/shrink and change in number depending on needs of cell

  
• Nuclear matrix


  Insoluble fibrillar-like protein network ('mesh') distributed throughout nucleoplasm, analogous to cytokeleton network in cytoplasm (composed of 3 major filament systems: microtubules, actin microfilaments & intermediate filaments), serves a structural role – maintains overall shape of nucleus, serves as a 'scaffold' – responsible for organizing nuclear subdomains and anchoring protein factors (e.g., proteins involved in DNA replication, transcription, RNA processing, etc)
• Nuclear envelope


  Separates the contents (e.g., genome) of nucleus from surrounding cytoplasm, serves as a barrier – requires regulated passage of molecules (e.g., RNA and proteins) between nucleus and cytoplasm, establishes unique composition of nucleus (compared to cytoplasm) and spatially regulates gene expression, provides structural framework for nucleus, composed of 3 main parts: nuclear membranes, nuclear lamina, nuclear pore complexes

  
• Nuclear membranes


  Inner and outer nuclear membranes - two concentric membranes (phospholipid bilayers)
• Scaffold


  Responsible for organizing nuclear subdomains and anchoring protein factors (e.g., proteins involved in DNA replication, transcription, RNA processing, etc)
• Very little is known about composition and assembly/disassembly of nuclear matrix