Cytologia i Biologia Komórki - Notatki z wykładu

Biological Membranes and Their Components

Biological membranes contain specialized areas known as lipid rafts, which are particularly rich in cholesterol and sphingolipids. These rafts exhibit a slightly lower fluidity than the rest of the membrane. The components of these membranes serve specific roles in maintaining cellular integrity. Cholesterol acts to limit the movements of phospholipids and provides stability to the membrane structure. Glycolipids are found exclusively in the outer layer of the bilayer in the plasmalemma (the membrane surrounding the cell), where they participate in the formation of the glycocalyx. Proteins within the membrane must be amphipathic and are categorized into surface proteins, which can be detached via mild extraction, and integral proteins, which cannot be separated without the disintegration of the phospholipid bilayer. These proteins perform an immense variety of functions as noted by the speaker. Finally, membrane carbohydrates are present in the form of either glycolipids or glycoproteins.

Mechanisms of Membrane Transport

There are three primary types of transport through biological membranes. Simple transport, or simple diffusion, occurs according to the concentration gradient and requires no energy expenditure. This process involves particles that can pass through the phospholipid bilayer, such as organic solvents like alcohols and benzene, steroid hormones, urea, and gas molecules like oxygen ($O_2$), nitrogen ($N_2$), and carbon dioxide ($CO_2$). Facilitated transport also moves according to the concentration gradient without requiring energy, but it applies to particles unable to penetrate the membrane spontaneously or those that do so in very small quantities; for instance, water requires aquaporins to pass in sufficient amounts. Examples of substances using facilitated transport include amino acids, sugars, and ions. This transport is mediated by transport proteins, which are divided into channel proteins and carrier proteins.

Channel proteins are tubular structures with a hydrophobic exterior that contacts the membrane and a hydrophilic interior that creates an environment for hydrophilic molecules to pass. These channels can be permanently open or change their permeability based on specific stimuli. There are ligand-gated channels, which open upon the binding of a signal molecule and remain open until the molecule dissociates. Mechanically opened channels are triggered by physical stimuli, such as the movement of stereocilia in the hair cells of the inner ear. G-protein-opened channels are activated, for example, by the stimulation of the muscarinic acetylcholine receptor in the heart muscle. Additionally, some channels are opened by changes in membrane potential, notably the sodium channels involved in nerve impulse transmission. Carrier proteins bind a molecule on one side and release it on the other, involving a change in the protein's conformation that returns to its original shape after the molecule dissociates. These include uniport (transport of a single substance), symport (transport of two substances in the same direction), and antiport (where the transport of one substance requires the simultaneous transport of another in the opposite direction).

Active Transport and Vesicular Movement

Active transport occurs against the concentration gradient and requires energy. Primary active transport uses energy directly in the form of ATP, such as the sodium-potassium pump ($Na^+/K^+$ pump), which the speaker notes is discussed so frequently in academic contexts that they decline to transcribe its full mechanism. Secondary active transport uses energy indirectly, relying on an electrochemical gradient generated by previous ATP consumption, such as the $H^+$ gradient in the mitochondrial intermembrane space or glucose transport powered by a sodium ion gradient. Endocytosis and exocytosis involve the fusion of membrane fragments to form vesicles around the substances being moved. Endocytosis, an active form of transport into the cell, is divided into pinocytosis (transporting surrounding fluid and dissolved substances) and phagocytosis (transporting large solids via specialized cells like Macrophages, granulocytes, Sertoliego cells, and retinal pigment cells). A variation of pinocytosis called transcytosis allows vesicles to pass through the cytoplasm unchanged to release their contents on the opposite surface, as seen in endothelial cells. Pinosomes form continuously in most cells. During phagocytosis, substances are surrounded by cytoplasmic protrusions that fuse to form fagosomy (phagocytic vesicles).

Selective uptake can also occur via receptor-mediated endocytosis. Ligand binding causes the formation of depressions, most of which are coated with the protein clathrin and are termed coated pits. These pits lead to the creation of endosomes containing the receptor-ligand complex, which dissociates in an acidic environment. Receptors then return to the membrane through specialized vesicles in a process called receptor recycling. Exocytosis is the secretion of vesicles containing substances into the external environment and can be constitutive exocytosis (continuous and independent of external factors) or regulated exocytosis (occurring in response to a specific factor, such as a hormone).

Intercellular Communication and Receptor Classes

Receptors are categorized into two main classes: intracellular receptors and membrane receptors. Intracellular receptors bind signal molecules capable of crossing the lipid bilayer, including steroid hormones, thyroid hormones, and Vitamin A derivatives. Their primary effect is the regulation of gene transcription. Membrane receptors appear on the cell surface, are mostly glycoproteins, and bind signal molecules that cannot penetrate the cell through simple transport. There are four classes of membrane receptors based on structure and signal transduction. These include receptors associated with channel proteins, where ligand binding causes a temporary opening of the channel. Catalytic receptors are transmembrane proteins whose cytoplasmic domain has catalytic activity, typically that of a protein kinase like tyrosine kinase; examples include EGF, PDGF, and insulin receptors. Multimeric transmembrane cytokine receptors bind cytokines. Finally, G-protein-coupled receptors use G-protein (a regulatory protein binding GTP) to trigger a cascade of secondary messengers, most commonly $cAMP$ or $Ca^{2+}$.

The Cytoplasm and Cytoskeletal Elements

The cytoplasm consists of the basic cytoplasm (cytosol), organelles, filamentous structures, and cytoplasmic inclusions. The cytosol exists as a highly hydrated gel containing proteins, metabolites, and nutrients. The cell skeleton, or cytoskeleton, includes three main components. Microtubules are $25\,nm$ in diameter and are built from heterodimers of alpha-tubulin and beta-tubulin. These polimerize in the presence of GTP into protofilaments, with $13$ protofilaments forming a single microtubule tube. They originate in the MTOC (microtubule organizing center) near the centrioles and undergo constant remodeling. Microtubules are polarized structures with a positive ($+$) pole, where polymerization dominates, and a negative ($-$) pole, where depolymerization dominates, leading to a state of dynamic instability. Tubulin dimers detach when their bound GTP is lost; polymerization requires a high local concentration of free tubulin and low levels of $Ca^{2+}$. Microtubules determine cell shape, organelle distribution (especially the ER and Golgi apparatus), and build cilia, flagella, and mitotic spindles. MAP (microtubule-associated proteins) stabilize microtubules and facilitate ATP-dependent transport of membrane-bound elements. Transport proteins include kinezyna (transporting toward the positive pole, or centrifugally) and dyneina (transporting toward the negative pole, or centripetally). Drugs such as kolchicyna, winblastyna, and winkrystyna inhibit the polymerization of mitotic spindle microtubules and are used as cytostatics in cancer therapy. Centrioles consist of nine triplets of microtubules arranged concentrically and linked by fibrillar proteins. Two pairs of centrioles form a centrosome, which initiates microtubule polymerization and, during mitosis, forms the division apparatus. Centrioles also form the basal bodies of cilia and flagella.

Microfilaments (actin filaments) are $6\,nm$ in diameter and consist of two intertwined chains of F-actin (polymers of G-actin). They are polarized and exhibit dynamic instability like microtubules. Polymerization depends on free actin and $Ca^{2+}$ and is initiated by ATP binding to actin; ATP hydrolysis causes depolymerization. Several proteins interact with microfilaments: Spektryna and ankiryna link them to integrins to support the membrane; Filamina networks the filaments to create the cortical layer of the cytoplasm; Żelsolina fragments the networks to facilitate shape changes and movement like endocytosis and exocytosis; motor proteins like miozyna I (pseudopod movement), miozyna V (intracellular transport), and miozyna II (muscle contraction) enable various forms of mobility; and Fibryna, alfa-aktynina, and fascyna cross-link filaments into parallel bundles for microvilli and stereocilia.

Intermediate Filaments and Ribosomes

Intermediate filaments are $10\,nm$ in diameter and composed of fibrous proteins with diverse amino acid compositions. They have globular domains at both ends, and their fibrous domains aggregate into dimers, tetramers, and octamers. The aggregation of eight tetramers creates a cord-like structure. Unlike other cytoskeletal elements, these are permanent structures. They form networks surrounding the nucleus or peripheral bundles linked to transmembrane proteins at cell junctions, providing cells and their protrusions with shape and mechanical strength. They form the nuclear lamina (lamins) and strengthen hemidesmosomes via plakiny. These filaments are tissue-specific: keratin filaments are specific to epithelial cells (cytokeratins type I and II); vimentin and vimentin-like filaments are found in mesenchymal tissues like connective tissue; desmin filaments (made of desmina and vimentin) appear in smooth muscle and skeletal muscle Z-lines; gliofilaments (GFAP and vimentin) are found in astrocytes; neurofilaments (NF-L, NF-M, NF-H, and alpha-internexin) occur in axons and dendrites; and nuclear lamins ($A, C, B1, B2, B3$) exist in all nucleated cells as attachment points for heterochromatin.

Ribosomes are structures found in the cytoplasm and on the rough endoplasmic reticulum composed of proteins and rRNA. They consist of a small ($40S$) and a large ($60S$) subunit, which together form an $80S$ ribosome. Areas rich in ribosomes stain basophilically and are called ergastoplazma. In protein synthesis, the small subunit matches aminoacylo-tRNA to codons while the large subunit creates peptide bonds. If the initial part of the peptide contains a signal sequence, the cytoplasmic ribosome attaches to the endoplasmic reticulum; otherwise, synthesis remains in the cytoplasm. Proteins for internal cell use are synthesized by cytoplasmic ribosomes, while those synthesized on ER-bound ribosomes are destined for "export."

Endoplasmic Reticulum and Golgi Apparatus

The Endoplasmic Reticulum (ER) is a system of membranes dividing the cell into compartments like tubules, vacuoles, and cisternae. The granular or rough ER (RER) consists of flattened cisternae covered in ribosomes. Its membrane contains protein complexes for ribosome attachment, including a docking protein (a membrane receptor recognizing the SRP protein-nucleic acid complex attached to the large subunit of the ribosome), ryboforyny (which bind the large subunit after SRP detaches), and enzymes like peptidazy and transferazy that modify the synthesized peptide. In the cisternae, the signal peptide is removed, and proteins undergo post-translational modifications. RER is abundant in cells intensive in protein synthesis, such as plasma cells or pancreatic cells. The smooth ER (SER) is a system of interconnected tubules in the peripheral cytoplasm. It is the site of lipid synthesis, certain stages of steroid hormone metabolism, glucose metabolism, and detoxification. It also segregates and modifies proteins from the RER and stores glycogen and lipids. It is abundant in steroid-producing cells. A specialized form, the sarcoplasmic reticulum, occurs in muscle fibers and stores calcium ions.

The Golgi apparatus is a membranous structure of parallel, flattened cisternae usually located near the nucleus. A group of $5$--$8 cisternae forms a diktiosom. Cisternae are arched and spaced $10$--$15\,nm apart. Those closer to the nucleus are cis cisternae (containing N-acetylglucosaminyltransferase), while those more distal are trans cisternae (containing galactosyltransferase and sialyltransferase), with intermediate cisternae between them. Numerous vesicles transport membranes between the ER, the cisternae, and the cell membrane, allowing the Golgi to participate in membrane remodeling and recycling. Vesicles also transport substances with the help of microtubules and dyneina (ER to cis pole), as well as kinezyna and miozyna II (both from the trans pole). Newly formed vesicles are "coated" by kotamerami (proteins that allow vesicles to bud off). The cis and intermediate cisternae also participate in modifying proteins and lipids.

Peroxisomes, Proteasomes, and Lysosomes

Peroxisomes are single-membrane vesicles containing approximately $50$ oxidative enzymes. Unlike lysosomes, they do not fuse with other vesicles. They are involved in inactivating toxic substances, breaking down hydrogen peroxide via katalaza, the beta-oxidation of long-chain fatty acids, and the synthesis of certain lipids and plasmalogens. They are numerous in cells active in detoxification and lipid metabolism. Proteasomes are cylindrical organelles built primarily of proteases. Proteins marked with ubiquitin undergo extra-lysosomal hydrolysis within them (typically damaged or faulty proteins). They prevent the accumulation of abnormal proteins, regulate the cell cycle by eliminating regulatory proteins, and break down certain antigens into peptides for presentation to the immune system.

Lysosomes are membrane-bound vesicles where the breakdown of macromolecular compounds occurs via hydrolytic enzymes at an acidic pH. They form through the fusion of hydrolase vesicles (primary lysosomes) from the Golgi apparatus with vesicles containing material for digestion. Their membrane contains proton pumps to maintain low pH. Secondary lysosomes include heterolizosomy (digesting extracellular material), autolizosomy (digesting the cell's own material), multivesicular bodies (digesting excess membranes), and residual bodies (remnants containing undigested material). Lysosomal enzymes belong to three groups: esterases (hydrolyzing ester bonds in fats, nucleic acids, or phosphate bonds in nucleotides; including nucleases and lipases), peptidases (hydrolyzing peptide bonds), and glycosidases (hydrolyzing glycosidic bonds). In some cases, like osteoclasts releasing collagenases, primary lysosomes release their contents outside the cell.

Mitochondria and Cytoplasmic Inclusions

Mitochondria function as the powerhouse of the cell. The speaker references the textbook author Zabla, noting that much of the mitochondrial information is standard knowledge but highlights specific key details. The inner membrane contains a high amount of cardiolipin, making it impermeable to small ions, which is essential for ATP synthase function. Within the matrix, mitochondrial grains (calcium and phosphorus deposits) are visible, as well as chaperones such as mit Hsp 70 and mit Hsp 60. Mitochondria are involved in apoptosis; the release of procaspase-2, -3, and -9, apoptosis-inducing factor (AIF), and cytochrome c from the matrix into the cytoplasm begins the apoptotic process. They are also responsible for steroidogenesis, as both mitochondrial membranes contain enzymes for synthesizing steroid hormones.

Cytoplasmic inclusions include glycogen, a storage material found as grains sized $25$--$30\,nm, which can be visualized with the PAS reaction. Lipids are high-energy storage materials found as non-membrane-bound droplets; in cells like the corpus luteum, they store cholesterol for steroid hormone production. Crystalline structures, such as those in the interstitial cells of the testis, consist of protein filaments and are thought to be a form of storage material. Pigment inclusions appear as grains in melanocytes, keratinocytes, and hair cells; in other cells like muscle or liver cells, they may appear as products of degeneration or aging, such as lipofuscic grains.

The Cell Nucleus and Nuclear Exchange

The cell nucleus contains almost all of the cell's genetic information and is characteristic of eukaryotic cells. While mostly mononucleated, there are binucleated cells (certain liver and cartilage cells), multinucleated cells (osteoclasts, bone marrow cells), and anucleated cells (erythrocytes, lens fibers). Nuclei are typically spherical or elongated, and can be segmented in monocytes and granulocytes. Active cells have large nuclei with loose chromatin, while inactive ones have small, dense nuclei. Nuclear size is greatest just before division and smallest after. The nucleus has three components: the nuclear envelope, chromatin, and the nucleolus.

The nuclear envelope consists of two membranes separated by a perinuclear space and is connected to the RER; the outer membrane is rough. The inner membrane is reinforced by the nuclear lamina (made of lamins), which provides shape and mechanical strength while organizing chromatin. Nuclear pores form where the inner and outer membranes join. These pores have an octagonal shape and, with nucleoporynami, form the nuclear pore complex composed of two peripheral rings (cytoplasmic and nuclear) and one central ring. Small molecules under $30\,kD$ diffuse freely, while larger ones require selective active transport. Proteins entering the nucleus must contain a nuclear localization signal (NLS), recognized by importins, transportins, and Ran proteins. Exported proteins contain a nuclear export signal (NES) recognized by exportins, while some proteins stay due to a nuclear retention signal (NRS).

Chromatin and the Nucleolus

Chromatin consists of a DNA double helix, various RNA particles, and proteins (histones and non-histones). Most nuclear DNA is repetitive DNA, including satellite DNA. Unique DNA builds the main fraction. Satellite DNA contains genes for tRNA and rRNA and aids in chromosome stabilization and crossing-over. The main fraction DNA contains sequences transcribed into hn RNA, which is processed into mRNA. Histones ($H1, H2A, H2B, H3, H4$) are basic proteins; their modification (methylation, phosphorylation, acetylation) affects chromatin packing and gene activity. Non-histone proteins include structural proteins (matriny, matrycyny, internal lamins), enzymatic proteins (nucleic acid metabolism), and regulatory proteins like transcription factors. The basic unit of organization is the nucleosome (DNA on a histone octamer). Nucleofilaments ($11\,nm$) form from linked nucleosomes, which then form a solenoid ($30\,nm$), the basic structure of interphase chromatin. Solenoids are packed into looped domains ($300\,nm$) by non-histone proteins. During prophase, these condense into chromatids ($700\,nm$). Chromosomes have short ($p$) and long ($q$) arms, light ($R$) and dark ($G$) bands. Kondensyny and Topoizomeraza II assist in condensation and preventing entanglement.

Euchromatin is less packed and transcriptionally active, while heterochromatyna is condensed and inactive. Heterochromatin is divided into constitutive (satellite DNA near centromeres and on the Y chromosome) and facultative (temporarily repressed, such as the Barra body or inactivated X chromosome). The nucleolus is a non-membrane-bound structure for rRNA synthesis and ribosome subunit formation. It disappears in prophase and reforms in telophase at the NOR (nucleolar organizing center). It has three parts: the fibrous center (rDNA, RNA polymerase I, SRP), the dense fibrous component (pre-rRNA, fibrylaryna, nukleolina), and the granular component (ribonucleoproteins, precursors of ribosome subunits). Nukleostemina and nukleolina assist in transporting precursors from the nucleolus.

Cell Cycle and Cell Division

The cell cycle is the period between divisions, consisting of interphase and the M-phase (mitosis). Interphase stages include $G1$ (anabolism and organelle recovery), $S$ (DNA replication doubling from $2c$ to $4c$, histone and centriole synthesis), and $G2$ (synthesis of tubulin and membrane components). The $G0$ phase is a resting stage where cells differentiate. The cycle involves cytoplasmic (Cdk activation), nuclear (DNA replication), and centrosomal (centriole duplication) cycles. Control is maintained by cyclin-dependent kinases (Cdks) and cyclins, which allow the cell to pass restriction points: transition from $G1$ to $S$ depends on the Cyclin $G1$ or Cyclin $D$--$Cdk2 complex; transition from $G2$ to mitosis depends on the mitotic Cyclin $B$--$Cdk2 complex (starting kinase). Mitosis ends with Cyclin $B$ degradation in proteasomes. Control is also exercised by anti-oncogenes like Rb and p53 proteins; loss of control leads to cancer.

Mitosis occurs in two stages: karyokinesis (division of genetic material) and cytokinesis (division of cytoplasm). The four stages are Prophase (condensation, nucleolus dispersion, centrosome migration), Metafaza (metaphase plate formation), Anafaza (centromere division and chromatid migration), and Telofaza (envelope restoration, spindle disintegration, decondensation). Meiosis consists of a reductional and an equational division, resulting in four haploid cells. Prophase I is prolonged and includes leptoten, zygoten, pachyten, diploten, and diakineza, where biwalenty form and crossing-over occurs.

Spindle Structure, Cytokinesis, and Cell Death

The mitotic spindle is a bipolar structure of microtubules and proteins. It includes kinetochore microtubules (linking centrioles to chromatids for migration via dyneina), polar microtubules (linking opposite centrioles and pushing them apart via kinezyna), and astral microtubules (anchoring the spindle to the cell cortex). Kohezyny prevent chromatid separation until anaphase; their malfunction causes nondisjunction and numerical chromosomal defects like aneuploidia. Cytokinesis begins in anaphase and ends after telophase, using a contractile ring of actin and miozyna II to create a division furrow. Nuclear envelope reorganization involves fragmentation and depolymerization via phosphorylation of lamins by Protein Kinase C and Cdk2. Reorganization involves Lap2, the lamin B receptor, and emeryna. Defosforylacja of lamin B allows the reconstruction of the nuclear lamina network. Mutations in these processes cause laminopatiami.

Cell death occurs primarily via necrosis or apoptosis. Necrosis is uncontrolled and results from strong damaging stimuli, leading to membrane disintegration, metabolic collapse, and a strong inflammatory response due to the release of cellular contents. Apoptosis is an ordered process of programmed cell death where the cell membrane remains intact. The cell turns into multiple apoptotic bodies that are phagocytosed by macrophages or neighbors, occurring during development without causing inflammation.