BIOCHEM 2.9 - CELLULAR HOMEOSTASIS AND ADDITIONAL DISORDERS

• The lecture is titled "Cellular Regulation and Case Presentations" for Block 2 Flipped Classroom.

Objectives

• Identify molecules and match their activity to steps in cell cycle regulation
• Examples include p53, pRb, p21, E2F and E2F promoter, CHK1 and CHK2, cyclins, CDKs, cyclin/CDK complexes, and INK4 family
• Assess the consequence for cellular division and/or molecular processes for changes in the following: binding at E2F promoter, Rb phosphorylation, cyclin availability, p53 activity, p21 activity, and INK4 family activity.
• Link changes in signaling with additional disorders; summarize features of disorders including cancer and proteus syndrome.
• Recognize clinical consequences for changes in cellular components.
• Connect the condition with the altered pathways/processes; identify clinical features of the condition.

G-Protein Coupled Signaling

• G-protein coupled signaling involves the activation of various downstream effectors.
• Phospholipase C activation leads to the production of inositol triphosphate and diacylglycerol (DAG).
• Inositol triphosphate increases calcium levels, activating calcium channels and nitric oxide synthase.
• Adenylate cyclase activation by GTP leads to cAMP production.
• cAMP activates protein kinase A (PKA).
• PKA phosphorylates CREB and allows translocation of the active subunit to the nucleus for transcription.

Pertussis Toxin

• Pertussis toxin alters cAMP activity by disrupting inhibitory regulation.
• Pertussis toxin (PTX) causes an increase in cAMP activity due to loss of inhibition/regulation.
• This leads to inhibition of AC activity and stimulation of cAMP-mediated signaling.

Cholera Toxin

• Cholera toxin alters cAMP activity by constitutive activation of adenylate cyclase.
• This leads to stimulation of cAMP-mediated signaling.

Proliferative Signaling

• Proliferative signaling in natural, regulated systems require signals for cell growth and division.
• This includes peptide hormones like insulin, growth factors such as epidermal growth factor, and cytokines like IL-6.
• Consequences of signaling include activation of transcription factors, growth/proliferation, and anti-apoptosis.
• AKT is a common mediator, linked to insulin, integrin, and PI3K.

Restriction Point

• The restriction point, classically known as the G1 checkpoint.
• After this point commitment to the cell cycle occurs, the cell commits to DNA replication
• Newer evidence questions the timing of no return but the steps associated are clear.
• Commitment to DNA replication involves hyperphosphorylation of Rb, activation of E2F-dependent transcription, and increased CDK2 activity.
• Hyperphosphorylation of Rb in cells re-entering the cell cycle from quiescence enables DNA synthesis and progression.

G1 to S Transition

• Mitogen activity/availability regulates cell division, and DNA damage/fidelity are critical regulators.
• Fatty acid metabolism regulates cell division.
• Lipid synthesis is required for cell division, conflicting evidence whether lipid checkpoint exists
• Fatty acid synthase (FASN) activity is connected to cell death.
• Inhibition of fatty acid metabolism in G1 inhibits entry into S phase.
• Fatty acid synthase blockers can prevent S phase entry, even when stimulated with growth factor.

Regulation of Cellular Division

• Control of cellular division is critical. Multiple steps include activating phosphorylation or inactivation.
• Checkpoint kinases include CHK1 and CHK2, increase p21 levels or by inhibiting CDC25 phosphatases.
• P53 is a DNA damage sensor.
• It acts as a transcription factor for p21, inhibiting cyclin D/CDK4 and preventing Rb phosphorylation.
• INK4 family has a p53-independent mechanism for G1 stalling in the presence of DNA damage.

Signal Mediators and Disease

• AKT is a common signal mediator.
• AKT genes are associated with cancer, cardiovascular disorders, diabetes, and neurological disorders.
• AKT1 overexpression is linked to cancer development and expression.
• AKT1 plays a role in cardioprotective processes that support growth and function.
• Pathogenic AKT1 variation can lead to tissue overgrowth.
• Proteus syndrome is a rare condition that can be caused by AKT1, characterized by overgrowth of bone, skin, and tissues.
• Growth in Proteus syndrome is asymmetrical.
• AKT1 mutations are common but not believed to be inherited; the condition develops due to mitotic error during early development.

Linking Human Disease

• The following conditions are a summary of the presentation: Ehlers-Danlos Syndromes, Duchenne muscular dystrophy, Rhabdomyolysis, Alport Syndrome, Laminins and Muscular Dystrophy, and ẞ-Thalassemia

Ehlers-Danlos Syndrome

• Ehlers-Danlos Syndrome is a group of 13 heritable connective tissue disorders, hypermobile EDS is common.
• Hallmarks are overly flexible joints and stretchy, fragile skin.
• Vascular Ehlers-Danlos syndrome is more severe, with of vascular/intestinal/uterine ruptures.
• Collagen and associated genes (affect collagen structure/function) are implicated.

Duchenne Muscular Dystrophy

• Duchenne muscular dystrophy is an X-linked recessive disorder more frequent in males than females.
• Incidence of one in 3600 live born males.
• It's the most common hereditary neuromuscular disease
• Causes progressive muscle degeneration.
• Symptoms appear prior to age six and life expectancy is reduced
• Therapeutics are centered around management of symptoms and improving quality of life
• Impacts the protein dystrophin, connection of intracellular with ECM

Rhabdomyolysis

• Damaged muscle releases proteins and electrolytes into blood.
• Marked creatine kinase elevation is observed.
• Mechanisms relate to ion movement, ROS, mitochondrial dysfunction, and lipid peroxidation.

Alport Syndrome

• Involves mutations in type IV collagen affecting production and deposition; 80% are COL4A5 on X chromosome.
• Impacts basement membrane in glomerulus, cochlea, and eye.
• Can weaken the basement membrane making it sensitive to proteolytic enzymes
• Significant cause of chronic kidney disease.

Laminins and Muscular Dystrophy

• Laminin-a2-related congenital muscular dystrophy (LAMA2-CMD) results in merosin deficient congenital muscular dystrophy type 1A (MDC1A).
• Mutations in LAMA2 gene (encodes laminin-a2 protein).
• Laminin-a2 is essential for assembly of laminin-211 and laminin-221.
• Disrupts communication between basal lamina and muscle membrane.
• Connection with integrins is affected.

Beta-Thalassemia

• Characterized by absent or markedly decreased accumulation of globin subunits of hemoglobin
• Beta-Thalassemias involve the beta-globin subunits.
• Over 200 different mutations have been identified.
• Mutations in beta subunits reduce their availability.
• Excess accumulation of alpha subunits occurs as they are unable to bind beta subunits.
• Homozygous-affected and compound heterozygous individuals have severe, hemolytic anemia.