Organelles and Movement of Materials Through the Cell

Lysosome

Digests obsolete organelles

Nucleolus

Synthesizes ribosomal RNA

smooth endoplasmic reticulum (SER)

detoxifies many types of drugs and foreign toxins

• the organelle primarily responsible for detoxifying many types of drugs and foreign toxins. This function is especially prominent in liver cells (hepatocytes), which contain a large amount of smooth ER. 

Golgi apparatus

Glycosylates proteins destined to be part of the cell’s glycocalyx

Rough ER

Synthesizes peptide proteins destined to be released from the
cell into the bloodstream

Smooth ER

Storage depot of calcium used in skeletal muscle contraction

Chaperone proteins

Facilitates proper folding of proteins

Golgi Apparatus

Fails to stain with H&E and Romanovsky stains, resulting in a
negative image (i.e. pale area in the cytoplasm)

Proteasome

Breaks down improperly folded proteins

Lysosome

Breaks down phagocytosed (ingested) infectious agents

Mitochondria

Organelle, other than the nucleus, that has its own DNA

Mitochondria

Production of ATP through fatty acid oxidation

True or false: organelles are metabolically active

True

• examples

  • nucleus

  • mitochondria

  • smooth & rough ER

  • golgi apparatus

  • lysosomes

  • cytoskeletal elements

What types of inclusions are metabolically inactive?

• fat

• glycogen

• melanin

• hemosiderin

• lipofuscin

Membrane Bound Organelles

•Lipid membranes separate different compartments                               of the cell, many containing mutually incompatible biochemical reactions.

What organelles have a double membrane?

• nucleus

• mitochondria

What organelles have a single membrane?

•Rough endoplasmic reticulum

•Smooth endoplasmic reticulum

•Transport vesicles

•Lysosomes

•Endosomes

•Peroxisome

•Golgi apparatus

Which organelles can be seen with visible light microscopy?

• nucleus

• golgi (negative image)

• nucleolus

***most organelles require electron microscopy to be seen

True or false: some organelles influence the color of the cytoplasm

True

•Nucleus, ribosomes (nucleic ACIDS):    Basophilic (blue)

•Mitochondria, Smooth ER:     Eosinophilic (pink).

*proteins are usually eosinophilic (pink)

What color does the nuclei appear to be with an H&E stain?

Basophilic (blue)

• negatively charged nucleic acids bind basic (usually blue/purple) dyes

What color does the nuclei appear to be with a Romanovsky stain (e.g. Wright Stain, DifQuik)?

Purple

• Cytoplasm is lightly basophilic or eosinophilic

True or false: histologic appearance of the nucleus can vary

True

•Usually round to oval

•Can be lobular or cigar shaped

— Nuclei vary in shape within cells, but will also be affected by plane of section – nuclei that are cigar shaped will appear round in a histologic section if cut in cross section.

What types of cells are multinucleated?

• osteoclasts

• skeletal muscle

• inflammatory macrophages

Heterochromatin

most highly condensed form of interphase chromatin in which genes cannot be expressed

• Used to silence genes

• Some DNA is permanently in heterochromatin (e.g. second X chromosome)

examples…

—> it is used to permanently inactivate one of the two X chromosomes in each female cell. 

—> It can also silence genes that encode or regulatory proteins important in early embryonic development but unnecessary in an adult cell.

Euchromatin

uncoiled chromatin with active DNA

• Most cell types express about 20 to 30% of their genes.  

Active

“____” cells tend to have more euchromatin than quiescent cells

• Transcribing a lot of DNA

• Making a lot of proteins

• Undergoing mitosis

less

More heterchromatin = a _____ (less/more) active cell

• highly condensed chromatin

Cell B

• B has a darker staining nucleus. This is probably a lymphocyte - the nucleus is much darker (i.e. more heterochromatin/more condensed chromatin) and it looks like there are some blood vessel cells in this area (endothelial cells). A is the nucleus of a hepatocyte (liver cell). These cells are very actively involved in protein synthesis — and therefore they need to continuously transcribe their DNA

terminally differentiated

Circulating neutrophils, eosinophils, and basophils are “____ _____”

•NOT capable of transcribing their DNA

•NOT capable of mitosis

***Most circulating WBCs have inactive nuclei (exception: monocytes & lymphocytes)

Which WBCs are capable of producing new proteins and capable of mitosis?

Monocytes and lymphocytes

True or false: Some cells can increase the amount of euchromatin when needed

True

• Most of these connective tissue cells (fibrocytes, black arrows) are quiescent, not actively producing connective tissue proteins and their nuclei contain a high proportion of heterochromatin.  If there is a wound, these cells can “wake up” and start producing the extracellular proteins that strengthen the tissue.  When this happens the chromatin unravels to increase the amount of euchromatin (fibroblasts, blue arrow).

mitosis

During ____, chromatin is looped and further condensed into a chromosome

• Chromatin is not only tightly condensed but is free - no longer contained within a nuclear envelope

nucleolus

• responsible for synthesis of ribosomes

• more prominent in cells actively involved in protein synthesis

• contains

  • rRNA

  • DNA w/ genes responsible for:

    - production & processing of rRNA

    - production of tRNA

• Ribosome subunits exported from nucleus and assemble in cytoplasm

Based on the nuclear features, which cell type is the most likely to be transcribing a lot of its DNA?

Cell type A appears more active

explanation…

Cell A:

• diffuse, fine granular chromatin (euchromatin; loosely condensed chromatin)

• prominent nucleolus

• lots of uncoiled DNA

• Active transcription = busy cell

Cell B:

• clumped, course granular chromatin

• mostly coiled DNA

• Can’t see a nucleolus

• low level transcription = inactive cell

• necrosis

• cell suicide

Cells die by 2 distinct mechanisms called ___ and ____

Necrosis

Cell murder

→ initiated by external factors such as hypothermia, excessive heat, low pH, microbial pathogens and ionizing radiation.

—> Cells that die by this process undergo swelling and membrane rupture. Necrosis always results in a secondary inflammatory response.

• mechanical injury

• hypoxia

• infectious agents

• toxins

Apoptosis

Cell suicide

→  involves internal programming to eliminate cells no longer needed or cells with irreparable internal damage.

• cells no longer needed

• irreparably damaged cells

• extrinsic & intrinsic signals

• TNFa(alpha)

Nuclear appearance in Cell Death (Necrosis vs Apoptosis)

Necrosis:

•Cells swell

•Nucleus swells

•Membrane dissolves

•Many cells involved

•Cells burst & contents trigger an inflammatory reaction

Apoptosis

•Cells shrink & cytoskeleton collapses

•Nuclear chromatin condenses (pyknosis & karyorrhexis)

•Nuclear envelope disassembles

•Cell forms blebs into fragments (apoptotic bodies)

•Macrophages “eat” them

—> microscopic appearance: fragments of dark nucleus, few dead cells, little to no inflammation; Because the cells are being eaten (phagocytized) and digested quickly, there usually few dead cells to be seen in a given area, even when large numbers of cells have died by apoptosis.

Which organelles is responsible for protein synthesis?

Ribosomes

• proteins that will remain in cytoplasm are synthesized by free ribosomes that float in cytoplasm

True or false: cells making lots of protein have large numbers of ribosomes which cause cytoplasm to appear basophilic (blue-purple)

True

• If a protein being made by the ribosome is destined for export it lands on the ER and passes out by Vesicle Transport. 

• If a protein intends to stay within the cell, ribosomes merely float around within the cell (polysome – chain of ribosomes on mRNA strand) and don't attach to the ER.

  • Ribosomes are stained more vividly by a Romanowsky stain than with H&E.

  

Protein folding is usually facilitated by ___

Chaperone proteins (aka heat shock proteins)

• Network of molecular chaperones promote efficient protein folding

• Several modes of action

• These increase in response to heat and attempt to refold denatured proteins

• Aberrantly folded proteins can cause aggregation → possible cellular damage.

What happens to proteins that fail to fold properly?

They are ubiquinated and then destroyed by a proteasome (an ATP-dependent protease)

• The proteolytic machinery of the cell and the chaperones compete with one another

Immature RBC contain ribosomes, while mature RBC do not.  Which of the indicated cells are immature RBC?

A is the immature RBC

• cells making lots of protein have large numbers of ribosomes which cause cytoplasm to appear basophilic (blue-purple)

What organelle can ribosomes be bound to?

rER

• Ribosomes bound to rER creates proteins for export to other organelles, to the plasma membrane, or out of body

• Each type of organelle has characteristic membrane & lumen proteins, phospholipids and other specialized molecules related to its function

• Ribosomes dock transiently on the rER

Free ribosomes vs ER-bound ribosomes

The main difference is their location and protein destination:

• free ribosomes in the cytoplasm make proteins for use within the cell

• while ribosomes bound to the rough endoplasmic reticulum (RER) synthesize proteins destined for export, secretion, or incorporation into cellular membranes, lysosomes, or other organelles. Ribsomes are released after synthesizing protein

  ***Structurally, they are identical, and a ribosome can switch between free and bound states depending on the protein it is synthesizing

What are the types of proteins produced in the rER?

Water soluble proteins (injected into lumen by ribosome)

• proteins are modified and folded

• packed into vesicles which are ultimately transported to Golgi (and beyond)

Transmembrane proteins (become embedded in rER membrane)

• destined for plasma membrane

  OR

• destined for membrane of endomembrane organelle

rER is well developed in ____cells

Secretory

Embedded ribosomes (RNA) cause what color tinge to cytoplasm

blue (especially w/ Romanovsky stains)

• Romanovsky stains, such as Wright stain, include a basic dye (Methylene Blue) which binds to nucleic acids and their phosphate groups.  These stains are very sensitive indicators of ribosomes and rER and almost all cell types will show some degree of cytoplasmic basophilia (i.e. some blue coloration) unless they are free of ribosomes and rER (e.g. mature erythrocytes). 

• Relatively quiescent cells will have pale blue cytoplasm, reflecting the low numbers of these organelles present.  A really deep blue color reflects a large number of ribosomes, either free or associated with rER – indicating active protein synthesis.

• Hematoxylin is not a true basic dye, so in sections stained with H&E, the basophilia associated with ribosomes and rER is not as pronounced.

Golgi Apparatus

• Organelle responsible for post-translational modification, sorting, and packaging

• Stack of  flattened saccules , near nucleus, without ribosomes

  • Usually curved – shaped dependent on cytoskeletal element and Golgi matrix proteins

• Proteins are modified as they move through the saccules becoming glycosylated (sugars added), phosphorylated (phosphates added), sulfated, etc. 

Golgi apparatus

Organelle that modifies proteins and generates the heterogeneous oligosaccharide strucutre seen in mature proteins

List the role of glycoproteins for protein function

• folding and stability

• interactions w/ other molecules

• cell signaling

• cell-cell communication

• cell protection, etc. (protection against proteosomes)

• part of glycocalyx on outer membrane

  • carbohydrate rich layer surrounding the cell membrane of many cells

    → aids in protection

    → permeability layer

    → aid in cell-cell communication/signaling

What are some of the destinations that the golgi sends vesicles to?

• secretory vesicles (e.g. collagen, hormones) (*material in them will be expelled*)

• components for plasma membrane (e.g. receptors)

• components of intracellular organelles (e.g. lysosomes)

True or false: golgi apparatus does not stain well with H&E or Romanovsky stains

True

• appears as a pale, perinuclear zone

• prominent in secretory cells

  • As a result, in secretory cells, you can sometimes see a negative image where the Golgi apparatus sits next to the nucleus.

  • Romanovsky(wright stain) = mostly membrane → methylene blue won’t stain

Vesicular transport

_____ _____ moves materials between organelles & the plasma membrane

Process of Vesicular transport

• Vesicles bud off from one organelle and fuse with another (or the plasma membrane) contributing

  • Proteins from rER (modified in Golgi)

  • Lipids from sER

• Things in lumen remain in lumen or pass into the extracellular space

• Vesicles move from place to place by motor proteins that pull them along the cytoskeletal proteins 

Vesicular transport

____ _____ allows movement of materials in & out of cell

Exocytosis

• outward secretory pathway

• Delivers new proteins, carbohydrates, and lipids to plasma membrane or the extracellular space

Endocytosis

• inward secretory pathway

• Plasma membrane components delivered to endosomes

• Components can be recycled or delivered to lysosomes for degradation

• Used to import various nutrients or ingest organisms or debris

Vesicular transport is facilitated by which proteins? Also describe process.

SNARE proteins

• Vesicle docks with the appropriate organelle (e.g. lysosome) through molecular markers on vesicle membrane and SNARE proteins

  • SNARE on vesicle (red line) interacts with SNARE on target (blue squiggles)

  • Holds vesicle in place

•SNARE proteins also catalyze vesicle fusion with membrane of target organelle or plasma membrane

How does the Clostridium botulinum neurotoxin cause botulism?

Acts on key SNARE proteins

• Synaptic vesicles containing neurotransmittors are released from neurons and stimulate muscles to contract. 

• Neurotoxin cleaves one of the SNARE proteins (exocytosis no longer works)

  • prevents fusion of vesicle

  • prevents neurotransmitter from stimulating muscle

• Result is muscle paralysis

• as toxin spreads, it can block nerves controlling the respiratory tract and heart, leading to death

How does exocytosis occur?

Through fusion of vesicle’s membrane with the cell’s membrane