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Cell Biology Basics

• Cell biology studies cells, including their structure, function, and behavior, requiring innovative techniques.
• The cell is the smallest unit of life, originating exclusively from pre-existing cells.
• Cells have the unique ability to reproduce themselves independently.
• Organelles cannot reproduce independently outside of a host cell.
• Viruses cannot reproduce without the host’s reproductive machinery.
• The cell is the fundamental building block of organisms, which then construct tissues, organs, and multicellular organisms.

Cell Size & Discovery

• Typical cell size ranges from 5 to 20 micrometers.
• Cells were discovered in the 17th century after the invention of the light microscope since they are invisible to the naked eye.
• Robert Hooke, using his light microscope in 1665, observed cells in a thin slice of cork and named them so because they reminded him of small rooms in a monastery.

Cellular Measurements

• 1 meter (m) is equal to 10^6 micrometers (µm) and 10^9 nanometers (nm), which is also equal to 10^10 angstroms (Å).

Basic Properties of Cells

• Life is the most basic property; cells grow and reproduce in culture.
• Cultured cells like HeLa (derived from Henrietta Lacks in 1951 by George Gey) are an essential tool for cell biologists.
• Cells are complex and organized, with highly regulated cellular processes.
• Similar structure and composition of cells among species remains conserved throughout evolution.
• Cells possess a genetic program, which is used to build each cell and the organism.
• Cells reproduce, and each daughter cell receives a complete set of genetic instructions.
• Cells acquire and utilize energy through activities such as photosynthesis and converting glucose into ATP.
• Cells carry out chemical reactions, the sum of which is called metabolism.
• Cells are able to engage in mechanical activities and respond to stimuli.
• Cells are capable of self-regulation and evolve over time

Prokaryotic vs. Eukaryotic Cells

• Prokaryotic and eukaryotic cells are the two basic types.
• Eukaryotes include protists, animals, plants, and fungi.
• Prokaryotes are all bacteria and emerged approximately 3.7 billion years ago.
• The distinction between prokaryotes and eukaryotes is size and type of organelles.
• All cells contain DNA, which is the genetic information store.
• Prokaryotic cells contain the Greek words "pro" for "before," and "karyon" referring to nucleus whereas eukaryotic cells derive their name from the Greek "eu," meaning "truly.”
• DNA is not segregated within a defined nucleus in prokaryotic cells, which is in contrast to that of eukaryotic cells.

Prokaryotic Cell Structure

• Prokaryotes feature a single compartment bounded by a membrane.
• The cytoplasm contains approximately 30,000 ribosomes, which account for the granular appearance.
• Contained within is the nucleoid which consists of a single circular DNA molecule, not separated by a membrane.

Common and Unique Features

• Common features of Eukaryotic and Prokaryotic cells include:
  • A similar plasma membrane
  • Genetic information in DNA with the same code
  • ATP to store energy
  • Shared metabolic pathways like glycolysis and the TCA cycle
  • Proteasomes for protein degradation of similar construction
• Unique features of Eukaryotic cells include:
  • A nuclear envelope separating the nucleus from the cytoplasm
  • Chromosomes that form into mitotic structures
  • Organelles bound by membranes
  • A cytoskeleton with motor proteins

Distinguishing Features

• Complexity: Prokaryotes are simple, whereas eukaryotes are functionally and structurally complex.
• Reproduction: Eukaryotes divide by mitosis, and prokaryotes use simple fission.
• Genetic Material: Prokaryotes have a nucleoid region; eukaryotes have a membrane-bound nucleus.
• Amount: Eukaryotes have more genetic material than prokaryotes.
• Form: Eukaryotes have many chromosomes consisting of DNA and histones with a single circular DNA and no histone proteins in prokaryotes.

Nucleus Structure

• The nucleus stores information in the cell and contains DNA.
• DNA is extremely long polymers that encode genetic instructions.
• The nucleus is surrounded by a double membrane called the nuclear envelope.
• Nuclear pores perforate the envelope and permit communication with the cytosol.
• The nuclear envelope is composed of inner and outer membranes.
• The nuclear lamina, made of fibrous network, offers structural support to the nucleus.
• Nuclear pore complexes are the only channels through which molecules travel to the cytoplasm.

Nuclear Lamina

• Supports the nuclear envelope and is made of lamins.
• The Integrity is regulated by phosphorylation/dephosphorylation.
• Mutations in Lamin A/C leads to Hutchinson-Gilford Progeria syndrome
• Lamin B mutation causes leukodystrophy (loss of myelin)
• Mutations in Lamin binding protein emerin causes Emery-Dreifuss muscular dystrophy (elbows, stiff neck and heels, and heart problems).

Trafficking of Molecules

• Traffic through nuclear pore complexes (NPCs):
  • DNA-binding proteins (histones, activators, repressors) are Imported.
  • Messenger RNA (mRNA)-binding proteins and ribosomal proteins.
  • Components of the nucleus (lamins).
  • Ribosomal proteins.
  • Shuttling nuclear transport receptors (importins) deliver proteins to the nucleus
  • mRNAs (with mRNA-binding proteins)
  • tRNAs (transfer RNAs)/40S and 60S ribosomal subunits (complexes of ribosomal RNAs [rRNAs] and ribosomal proteins) are Exported

Vertebrate Nuclear Pore

• A vertebrate nuclear pore complex (NPC) contains a scaffold anchoring it to the nuclear envelope, a cytoplasmic and nuclear ring, and nucleoplasmic filaments.
• It is a huge complex that is 15–30 times the mass of a ribosome and has octagonal symmetry with a Channel width of 20–30 nm.
• FG (phenylalanine-glycine) domains form a sieve that is hydrophobic and blocks diffusion of macromolecules larger than 40,000 Daltons.

GTP

• Binding of GTP (activation) needs a GEF (guanine nucleotide exchange factor).
• Hydrolysis of GTP to GDP (inactivation) needs GAP (GTPase activating protein).

Protein Import

• Proteins synthesized in the cytoplasm get targeted to the nucleus by an NLS (nuclear localization signal).
• Proteins with NLS bind to an NLS receptor (importin α/β heterodimer).
• The resulting protein/importin complex then associates with cytoplasmic filaments.
• Complex passes through the NPC.
• The importin complex associates with a GTPase called Ran.
• β complex is transported back to the cytoplasm where Ran gets modified and returns to the Nucleus which enables Importin α to be transported out of the nucleus via Exportin

ER and Ribosomes

• The outer nuclear membrane is continuous with the rough endoplasmic reticulum (ER).
• The space between the inner and outer nuclear membranes connects to the ER lumen.
• Eukaryotic (vertebrate) and prokaryotic ribosomes are comprised of rRNA and proteins that are of varying molecular weight.
• Ribosomes consist of larger and smaller subunits, the assembly is slightly different for each type.
• It is thought in early cell evolution that the nuclear envelope arose from the plasma membrane where the DNA and some membrane-bound ribosomes became enclosed.

Nucleolus and Chromatin

• The nucleolus, found in the nucleus, is a suborganelle for ribosome assembly.
• Ribosomal proteins enter the nucleus.
• These ribosomal proteins are sent to the nucleolus to bind to pre-rRNA (pre-ribosomal RNA).
• Pre-rRNA undergoes cleavage to produce multiple rRNAs.
• Ribosomal proteins then combine with rRNAs to make the 40S and 60S ribosomal subunits, which are then exported.
• Chromosomal DNA is organized into chromatin fibers combined with specialized proteins.

Chromatin Makeup

• Chromatin is composed of eukaryotic DNA and associated proteins.
• Chromatin is comprised of twice more protein than it is DNA.
• The major proteins are histones, small proteins (11 to 23 kDa) that contain arginine and lysine that facilitates to the negatively charged DNA molecule.
• There are 5 major types of histones including: H1, H2A, H2B, H3, H4.
• Histone protein content is similar across different eukaryotes.
• Chromatin contains almost an equal mass of nonhistone chromosomal proteins of more than 1000 types.

Nucleosomes

• The basic structural unit of chromatin is the nucleosome.
• DNA is wrapped around an octamer of histones which include H2A, H2B, H3 and H4.
• Linker DNA joins nucleosome core particles.
• Packaging DNA with nucleosomes produces a 1/7th compaction of the original DNA.
• Non-histone proteins interact by binding with linker DNA.

Higher Order Compaction

• Further packaging occurs through histone interaction and results in 30nm fibers.
• These zigzag or solenoid structures compact the DNA ~6x fold
• Subsequently the 30 nm fibers are looped into 80-100 nm supercoiled loops secured by cohesin protein.
• Mitotic chromosomes demonstrate maximal compactness with a ratio of about 10,000:1, 1 µm of chromosome length contains 1 cm on DNA.
• Chromatin condenses into chromosomes during cell division, which are easily seen with a microscope.

Heterochromatin vs. Euchromatin

• Euchromatin returns to a dispersed state after mitosis.
• Heterochromatin persists in a condensed state during interphase.
• Constitutive heterochromatin remains condensed all the time and it is mostly around centromeres and telomeres. Consists of highly repeated sequences and few genes.
• Facultative heterochromatin becomes inactivated during certain development during phases, like X-inactivation.

Mitochondria

• Mitochondria play a role in generating metabolic energy in eukaryotic cells by oxidizing carbohydrates and lipids into ATP through oxidative phosphorylation.
• ATP is used in a range of energy reaction within cells.
• Mitochondria consumes oxygen and releases carbon dioxide which is used for cellular respiration
• Mitochondria can appear as a branched, interconnected tubular network.
• Balance between fusion and fission determines mitochondrial morphology.

Mitochondrial Formation

• Single mitochondrion is about ~4 µm long.
• Dynamic organelles undergo dramatic changes and can can with one another (fusion) or split into two (fission).
• Fission is induced by contact with endoplasmic reticulum (ER) tubules.
• Mitochondria arise from preexisting mitochondria which divide via fission.

Mitochondrial Membranes and Structure

• Mitochondria have an inner and outer membrane.
• The intermembrane space is between the inner and outer membranes
• The inner membrane is highly folded (cristae), with it extending into the interior (matrix).
• The matrix contains hundreds of types of enzymes alongside circular DNA molecules (mitochondrial genome) and some special mitochondrial ribosomes.
• Inner mitochondrial membrane the ATP synthase site.
• The Outer membrane contains enzymes that convert lipid substrates to forms that are metabolized in the matrix.

ATP Production

• Electron transfer generates a proton gradient across the inner membrane, which drives ATP production by the ATP synthase.
• High-energy electrons taken from NADH and FADH2 are passed along the electron-transport chain in the inner membrane to oxygen (O2) in the process known as oxidative phosphorylation.
• Pyruvate and fatty acids are broken down to acetyl CoA, and that are then metabolized by the citric acid cycle, which produces NADH and FADH2
• This e- transport generates a proton gradient which makes ATP!

Electron Transfer

• The five types of electron-transfer carriers include:
  • Flavoproteins (contain NAD+ and FAD+ )
  • Cytochromes (contain heme such as Fe3+ → Fe2+)
  • Copper-containing proteins (Cu2+ → Cu1+)
  • Ubiquinone, which carries but isn't associated with a protein
  • Iron-Sulfur proteins

Glycolysis

• Glucose is phosphorylated and phosphorylated again.
• The six-carbon bisphosphate is split into two three-carbon monophosphates.
• The three-carbon aldehyde is oxidized into the coenzyme NAD+ to NADH and phosphorylated to form an acyl phosphate
• The phosphate group from C1 is transferred to ADP to ATP by molecule oxidized, 2 per molecule oxidized
• This substrate gets rearranged and dehydrated
• Generated ketone gets transferred to ADP and is a substrate-level phosphorylation
• Two ATPs are formed per glucose oxidized.

Pyruvate

• In the presence of oxygen, pyruvate goes through oxidative phosphorylation to make ATP or fermentation.
• Fermentation regenerates NAD+ and allows cells to metabolize glucose and produce small amounts of ATP

TCA cycle

• The TCA cycle occurs with acetyl CoA is condensed with oxaloacetate to produces citrate.
• Then, two carbons are oxidized out in the form of CO2, and oxaloacetate is regenerated.

Mitochondria Production Summary

• Each pyruvate generates 4 NADH with 3 from the cycle and one from the AcCoA production, it also generates FADH2 with 2 molecules of GTP The total amounts produced include 10 NADH 2 FADH2 2 molecules of GTP
• Each NADH makes ~2.5 ATP, each FADH2 makes ~1.5 ATP, and about 30 ATP from a single glucose molecule.

Electron Transport Chain

• 5 complexes participate in the electron transport chain including; Complex I (NADH dehydrogenase), complex II, complex III, the complexes have several proteins attached through the mtDNA or nDNA.
• Electrons from NADH or FADH2 get passed to ubiquinone
• Electrons then pass from coenzyme Q to complex III.
• Lastly, electrons are all transferred to chain which carries electrons to a peripheral membrane
• Then, they are transferred to cytochrome c and form complex IV or to cytochrome oxidase.
• Complex IV transfers the electrons to O2 and makes water.

Proton Transfers

• Electron transfers generate energy that pumps protons from the matrix to the inner membrane and form the lumen of the intermembrane which drives ATP synths-

Blocking ATP

• ATP production can be blocked by poisons like cyanide and some metals that bind to the catalysis sites of complex 4
• Endogenous proteins can also uncouple it from ATP

Mitochondria Disorders

• Can affect muscle and nerve tissue because of a higher demand for ATP, and this creates ragged-red fibers
• A buildup of ROS creates mutations
• mtDNA mutations can form adult orders like PD.
• Heroin contaminated with drugs can also result in complex 1 activity
• petite colonies also come from mitochondrial disorders and have different sources to regenerate energy

Mitochondrial Origins

• Mitochondria are thought to come from endosymbiont theory including chloroplast and are derived from prokaryotic cells

Evidences for Endosymbiosis

• Outer membrane of bacteria and mitochondria contain porins
• Inner membrane of both contain lipid cardiolipin
• Both mitochondria divide from fission and the single circular DNA 5. Ribosomes are the same and use 70s

Chloroplasts

• Chloroplasts have an outer and inner membrane as well as the stroma which is analogous to mitochondria matrix and thylakoids for stacking

Plant Cells

• The largest characteristic organelle are found in plant cells as photosynthetic for light reactions that make molecules from organic dioxide and use ATP and NADPH to switch CO2.
• The photosynthetic unit contains hundred of chlorophyll molecules; the reactions enter transfers electrons with transfer and antennae to transfer those photons to the reaction center.
• photosynthetic units found in PSI and PSII are complexes embedded in the thylakoid membrane

Light Harvesting

• After a center reaction happens for PSII with the water breaking down O2, protons are released and the electrons enter the transfer chain to be transferred to complex.
• They then transfer electrons to cytochrome c and form complex IV or to cytochrome oxidase.
• This e- transport generates a proton gradient which makes ATP.

Chloroplasts in Detail

• Pigments within the plant capture light
• 2H2O is split because it's water splitting enzymes to 02 and 4H
• the protons travel in the plastoquinome and H, H
• then in the Cytochrome
• lastly in photosystem 1 when the proton levels increase and use ferredoxin to combine and to generate a system that adds ADP and protons that make to a system that makes ATP +H

Thylakoids Details

• Electrons transferred from PS11 to Cytochrome
• then combine and to generate a system that adds ADP and protons which make to a system that makes ATP

Metabolic Differences

• Both mitochondria and chloroplasts: Generate ATP and use a H+ pump, have DNA and ribosomes, and have two membranes
• Only chloroplasts have an additional membrane and Cristea for the mitochondrial cell
• Terminal electron acceptor in OP is O2 or NADP+
• OP requires O2 but PSLR utilize CO

Carbon Cycle

• Six of carbon dioxide then run rubisco
• Rubisco then makes carboxylase intermediate
• It then splits into PGA and 6 RuBP
• then the carbon cycles