Ultrastructure of cells

Nihar Ranjan Jana

School of Bio Science
Email: nihar@iitkgp.ac.in
BS20001
 Microscopes
Microscopes are used to observe small
objects invisible to the naked eye.
The quality of the image depends on:
 Magnification: the microscope’s power to
increase an object’s apparent size
 Resolution: minimum distance between two
distinguishable points
 Contrast: visible differences between different
parts of the sample
Up to 0.2 μm
Microscope Contrast Specimen Contrast
Bright field

By selective and differential staining of

cellular components

Differential Interference Contrast

Phase contrast

Dark field
 Electron Microscopy
Subcellular structures are studied by electron microscopes.
They are of two types:

• Scanning Electron Microscopes (SEM) focus a beam of

electrons onto the surface of the sample and provide
images that give 3D representation of the sample. SEM is
used to study surface structure of objects

• Transmission Electron Microscopes (TEM) focus a beam

of electrons through the sample. TEMs are used to study
the internal structure of the cell
Electron Microscopy: SEM vs. TEM
Scanning Electron Microscope (SEM) Transmission Electron Microscope (TEM)
Electron Microscopy: SEM vs. TEM
Scanning Electron Microscope (SEM) Transmission Electron Microscope (TEM)

Surface features Intracellular ultrastructure

Fluorescence Microscope
 Confocal microscope
• A pinhole focuses the illumination at a point
• Another pinhole collects emitted light (signal) only from a point (focus)
• Noise from out of focus points in specimen is excluded

Point of focus of illuminating light

and emitted light are same
Why are Cells Small?
 All cells are prokaryotic or eukaryotic

• Prokaryote: single-celled organisms,

and all are bacteria.
• Eukaryote: single-celled or multi-
cellular organisms

• Pro = before
• Eu = true
• Karyon = nucleus
Prokaryotic cells have a simpler internal organization than eukaryotic cells

Similarities:
1. Plasma membrane
2. Cytoplasm
3. Genetic material (DNA)
4. Energy currency
5. Enzymes and coenzymes
 Differences:
Prokaryotic cells Eukaryotic cells
No membrane-enclosed Have well defined nucleus (membrane-
intracellular compartment to enclosed intracellular compartment) to
house genetic material (DNA) house genetic material (DNA)
Prokaryotic cells lack most of the Eukaryotic cells have well defined and
complex membrane bound complex membrane bound internal
internal organelles organelles
Prokaryotic cells have a single Eukaryotic cells have paired
circular chromosome chromosomes
Prokaryotic cells lack histone
Eukaryotic cells have histone proteins
proteins;
Plant and fungal cells have both
Prokaryotic cell wall has
cellulose and chitin in cell wall. No such
peptidoglycan;
cell wall in animal cells

Prokaryotic cells
No membrane-enclosed
intracellular compartment to
house genetic material (DNA)
Prokaryotic cells lack most of the Eukaryotic cells have well defined and
complex membrane bound
internal organelles
Prokaryotic cells have a single
circular chromosome
Prokaryotic cells lack histone
proteins;
Prokaryotic cell wall has
peptidoglycan;
Differences:
Eukaryotic cells
Have well defined nucleus (membrane-
enclosed intracellular compartment) to
house genetic material (DNA)
complex membrane bound internal
organelles
Eukaryotic cells have paired
chromosomes
Eukaryotic cells have histone proteins
Plant and fungal cells have both
cellulose and chitin in cell wall. No such
cell wall in animal cells

http://igbiologyy.blogspot.in/2014/03/chromosomes-dna-genes-and-alleles.html
 Differences:
Prokaryotic cells Eukaryotic cells
No membrane-enclosed Have well defined nucleus (membrane-
intracellular compartment to enclosed intracellular compartment) to
house genetic material (DNA) house genetic material (DNA)
Prokaryotic cells lack most of the Eukaryotic cells have well defined and
complex membrane bound complex membrane bound internal
internal organelles organelles
Prokaryotic cells have a single Eukaryotic cells have paired
circular chromosome chromosomes
Prokaryotic cells lack histone
Eukaryotic cells have histone proteins
proteins;
Plant and fungal cells have both
Prokaryotic cell wall has
cellulose and chitin in cell wall. No such
peptidoglycan;
cell wall in animal cells
Histone proteins
 Differences:
Prokaryotic cells Eukaryotic cells
No membrane-enclosed Have well defined nucleus (membrane-
intracellular compartment to enclosed intracellular compartment) to
house genetic material (DNA) house genetic material (DNA)
Prokaryotic cells lack most of the Eukaryotic cells have well defined and
complex membrane bound complex membrane bound internal
internal organelles organelles
Prokaryotic cells have a single Eukaryotic cells have paired
circular chromosome chromosomes
Prokaryotic cells lack histone
Eukaryotic cells have histone proteins
proteins;
Plant and fungal cells have both cellulose
Prokaryotic cell wall has
and chitin in cell wall. No such cell wall in
peptidoglycan;
animal cells
 Bacterial Cell Wall
• Lies outside the cell membrane in nearly all
bacteria (except mycoplasma and some
archaebacteria)

• Two important functions:

1. Maintains the characteristic shape

2. Prevents osmotic lysis

 Bacterial Peptidoglycan Layer

Gram +ve Gram –ve

S. aureus E. coli
Components of Bacterial Peptidoglycan Layer

(NAG) (NAM) (NAG)

Gram –ve bacteria like E. coli

Components of Bacterial Peptidoglycan Layer
 Gram Staining

A Gram stain of mixed Staphylococcus aureus (Gram

positive cocci) and Escherichia coli (Gram negative bacilli),
the most common Gram stain reference bacteria
 Controlling Bacteria by
Damaging Cell Walls

• The antibiotic penicillin blocks the final stages of

peptidoglycan synthesis

• The enzyme lysozyme, found in tears and other

human body secretions, digests peptidoglycan
 Cell Membrane

• Structure: phospholipid bilayer with proteins that function as

channels, markers, and receptors. Also contains cholesterol
which provides rigidity.
• Function: selectively permeable boundary between the cell
and the external environment.
 Bacterial Internal Structures
Bacterial cells typically contain (in their
cytoplasm):
Ribosomes
Nucleoid region
Vacuoles
Certain bacteria sometimes
contain endospores
 Prokaryotic Nuclear Region (Nucleoid)

This centrally located nuclear region consists mainly of DNA

(deoxyribo nucleic acid), but also contains RNA (ribonucleic
acid) and protein

DNA: Usually one large, circular chromosome (in Escherichia

coli DNA circle measures ~1400 µm)
The DNA is coiled extensively with RNA and
nucleoid proteins

Plasmids: Extrachromosomal pieces of smaller, circular DNA

 WHAT IS A VIRUS?

 A virus is a submicroscopic particle

 A virus is an infectious, obligate, intracellular parasite

 Contains only nucleic acids as genomic material and proteins coat to

protect its genome

 Viral genome can be DNA or RNA

 Lacks ribosomes or any membrane bound organelles

 Viral genome directs synthesis of viral components using host-cell

machinery
 Are viruses are alive?

A virus is an organism with two phases

Nonliving phase living phase

Virion
(Polio virus)
Polio virus infected cell
 Baltimore classification of virus

Parvovirus

Retrovirus (HIV)

Adeno virus
Herpes simplex virus 1. ds DNA
2. ss DNA
3. ds RNA
4. ss (-) RNA
5. ss (+) RNA
Reovirus 6. Ss (+) RNA with
Rotavirus DNA
Poliovirus intermediate
Dengue virus

Influenza virus
Ebola virus
Rabies Virus
 Flow of Genetic Information: Updates

RNA Replication

Transcription Translation
Reverse
Replication Transcription
DNA RNA Protein

David Baltimore
Nobel Prize in 1975
 Virus growth
Poliovirus (+ve) sense RNA virus

Steps of virus life cycle:

1. Attachment
2. Entry
3. Uncoating/ Disassembly
4. Translation
5. Replication
6. Assembly and release
 Virus growth

Viruses replicates by
assembly of preformed
components into many
particles

Make the parts and assemble into

final product
 How to measure virus titer

Plaque assay for animal viruses:

• Developed by Renato Dalbecco in 1952; Nobel prize in 1975

• Depends upon the cytopathic effect caused by the virus into a monolayer
of cells
• Gives the measure of infectious virus units per milliliter of sample
• Does not account for the noninfectious or defective virus particles
Plaque assay
 Equilibrium and non-equilibrium viruses

Equilibrium viruses are have been long term parasites for the host species

They are usually non-lethal but spreads well

Example: common cold

Non-equilibrium viruses have recently jumped from another species

They are some times lethal, may spread poorly or well

Represent most of the deadly viruses

Equilibrium and non-equilibrium viruses

Equilibrium viruses Non-Equilibrium viruses

Polio Influenza (Birds)

Common cold HIV (Chimps)
Measles SARS(Bats?)
Herpes Ebola (Bats)
Hantaan (Rodents)
 Human Immunodeficiency Virus (HIV)

• First appeared in humans in early 1980’s

• Classic non-equilibrium virus

• Endemic in African monkeys and jumped into

humans around 90 years ago

• It infects and kills helper T cells and and hence

wipes out a major component of our immune system

• Gets integrated in our genome – infected person

becomes a life long host for the virus
Hijacking the host - HIV
 Hijacking the host - HIV

• Recognizes specific CD4 receptor found in the T helper cells

• Post entry viral enzyme. The reverse transcriptase performs the reverse
transcription process

• Synthesized the proviral DNA

• Transported to the nucleus of the host cells

• Another viral enzyme, the Integrase integrates the proviral DNA into the
host genome – establishment of latency

• The provirus can express viral proteins and RNA using host machinery
to produce more virus – lytic cycle
Ultrastructure of
Eukaryotic cells
 Cytoplasm

• Structure: gelatin-like fluid that lies inside the cell membrane

• Function:
-contains salts, minerals and organic molecules
-surrounds the organelles

Cytoplasm
 Nucleus
• Structure: the nucleus is a compartment that contains another
compartment called a nucleolus
• Function:
-storage center of cell’s DNA
-manages cell functions
Nucleus
 Endoplasmic Reticulum (ER)

 Structure: a system of membranous

tubules and sacs

 Two types:
• Rough Endoplasmic Reticulum-
ribosomes attached on the
membrane
• Smooth Endoplasmic Reticulum-
no ribosome attached

 Function:
 Part of intercellular highway (a path along which molecules move from one
part of the cell to another)
 Manufactures, processes, and transports a wide variety of biochemical
compounds (lipids, proteins) for use inside and outside of the cell.
 Intracellular store of Ca2+ ions that is used in cell signaling responses
Rough and smooth ER

A. An electron micrograph
showing the rough ER in a
dog’s pancreatic cell that
makes and secretes large
amounts of digestive
enzymes. The cytosol is
filled with closely packed
sheets of ER, studded with
ribosomes

B. Electron micrograph of a
Leydig cell in the human
testis that secretes
testosterone (steroid
hormone), showing
abundant smooth ER
 Golgi Apparatus
• Structure: stacked flat sacs
• Function: receives proteins from
the rER and distributes them to
other organelles or out of the cell
(receiving, processing,
packaging, and shipping)
Mitochondria and Chloroplast
Lysosome
Cytoskeleton: Support, Motility and Regulation

• Cytoskeleton helps maintain correct shape and proper

internal structures of cells

• It interacts with motor proteins to assist in motility

• It helps cells to change shape and move from one place

to another

• During mitosis, it pulls the chromosomes apart and

helps in cell division
 Three types of protein fibres constitute cytoskeleton
1. Microfilaments (Actin filaments) determine cell shape and drive whole-cell
locomotion.
2. Microtubules determine the positions of cellular organelles and direct intra-
cellular transport.
3. Intermediate filaments provide mechanical strength.
Localization
Actin filaments or microfilaments
• Actin exists as a globular
monomeric protein called G-
actin and as a filamentous
polymer called F-actin, which
is a linear chain of G-actin.
• In the filaments, the subunits
are organized as a tightly
wound helix.
• All subunits in an actin
filament point toward the
same end of the filament.
Consequently, a filament
exhibits polarity; that is, one
end differs from the other.
 Each type of cytoskeletal filament is
constructed from smaller protein subunits
 Cellular locomotion
Forces generated in the actin-filament-rich cortex help move a cell forward:
• Protrusion: Actin polymerization at
the leading edge of the cell pushes
the plasma membrane forward
(protrusion) and forms new regions of
actin cortex (red).
• Attachment: New points of
anchorage are made between the
bottom of the cell and the surface
(substratum) on which the cell is
crawling.
• Contraction at the rear of the cell—
mediated by myosin motor proteins
moving along actin filaments—then
draws the body of the cell forward.
• New anchorage points are
established at the front, and old ones
are released at the back, as the cell
crawls forward. The same cycle is
repeated over and over again,
moving the cell forward in a stepwise
fashion.
Actin filament polymerization at the leading edge
pushes lamellipodium forward
 Continuous actin nucleation at
the plus end facing membrane
 Branching structures push the
plasma membrane forward
 Plus ends are protected from
depolymerizing by capping
proteins
 Minus ends nearer the center of
the cell disassemble through
depolymerization
 The web of actin as a whole
thereby undergoes a continual
rearward movement due to the
assembly of filaments at the
front and their disassembly at
the rear
Microtubule subunits: α and β tubulins
Microtubule associated Motor Proteins:
Kinesins and Dyneins

Kinesin

Dynein Kinesin and dynein mediated

transport of vesicles and
ll
 Books and resources

Video links:
https://www.youtube.com/watch?v=URUJD5NEXC8
https://www.youtube.com/watch?v=B_zD3NxSsD8

https://www.dropbox.com/s/qoseir2l5tn8jmk/Nihar-
Lecture5_Ultrastructure%20of%20Cells.ppt?dl=0