Lecture 30 – Receptor Protein-Tyrosine Kinases; Steroid Hormone Signalling

Q: What is convergence in signal transduction?

Convergence is when signals from unrelated receptors lead to the activation of a common effector. For example, GPCRs, RTKs, and integrins all bind different ligands but can all lead to a docking site for Grb2.

Q: What is divergence in signal transduction?

Divergence is when a single signal reaches a variety of different effectors. A single stimulus can send signals along multiple different pathways simultaneously.

Q: What is cross-talk in signal transduction?

Cross-talk is when signals are passed back and forth between different signalling pathways, allowing them to influence each other.

Q: How does convergence result in similar outcomes in different target cells?

Convergence can result in a similar set of growth-promoting genes being produced in different target cells, because different upstream receptors (GPCRs, RTKs, integrins) all funnel through a shared downstream component like Grb2.

Q: Give an example of cross-talk between two major signalling pathways.

cAMP-activated PKA blocks signals from Ras to Raf (MAPKKK), inhibiting MAPK activation. Additionally, Ca²⁺ and cAMP can influence each other's pathways.

Q: How can cAMP inhibit the MAP kinase cascade?

cAMP activates PKA, which phosphorylates and blocks the signal from Ras to Raf (MAPKKK), thereby inhibiting downstream MAPK activation.

Q: How can both PKA and Rsk-2 kinase converge on CREB?

Both PKA (from the GPCR/cAMP pathway) and Rsk-2 kinase (from the MAPK cascade) can phosphorylate the CREB transcription factor, which then binds CRE elements to activate gene expression — an example of convergence.

Q: What are steroid nuclear hormones? Give examples.

Steroid nuclear hormones are lipid-soluble signalling molecules that can cross the plasma membrane and bind intracellular receptors. Examples include glucocorticoids, progestins, estrogens, androgens, and mineralocorticoids.

Q: How do steroid hormones transduce signals to the nucleus?

Steroid hormones enter the cell, bind their receptor in the cytoplasm, causing release from chaperone proteins (e.g., HSP90), homodimerization of the receptor, and translocation into the nucleus where the receptor complex binds DNA response elements and recruits coactivators to activate target genes.

Q: What are the three functional domains of a steroid hormone transcription factor?

(1) A DNA-binding domain that binds to DNA response elements, (2) a trans-activation domain that activates transcription, and (3) a ligand-binding domain that binds the hormone to activate the transcription factor.

Q: What are Type I nuclear steroid hormone receptors and where are they found before activation?

Type I receptors include the androgen receptor, estrogen receptor, and progesterone receptor. Before activation, they are present in the cytoplasm bound to chaperone proteins such as HSP90.

Q: Describe the activation sequence of a Type I steroid hormone receptor.

Ligand binding causes release of the receptor from the HSP90 chaperone, followed by homodimerization, translocation into the nucleus, binding to a DNA response element, and association with transcriptional coactivators to activate target genes.

Q: What is the glucocorticoid receptor (GR)?

The GR is a nuclear receptor that includes a ligand-binding domain and functions as a DNA-binding transcription factor. It binds to glucocorticoid response elements (GREs) in DNA.

Q: What is a glucocorticoid response element (GRE)?

A GRE is a DNA sequence that the glucocorticoid receptor binds to. It is a palindrome, meaning both DNA strands have the same 5′ to 3′ sequence.

Q: What structural motif does the glucocorticoid receptor use to bind DNA?

The glucocorticoid receptor is a zinc-finger protein. Each zinc finger has a zinc ion coordinated between two cysteines and two histidines, and the fingers project independently into successive major grooves in the target DNA.

Q: What three types of chromatin modifications control chromatin state and gene regulation?

(1) DNA methylation, (2) histone acetylation, and (3) histone methylation.

Q: Describe the model for transcriptional activation at a promoter by the glucocorticoid receptor.

Active GR binds DNA and recruits the coactivator CBP. CBP acetylates nucleosome histones. The acetylated histones then recruit chromatin remodelers (SWI/SNF complex), which disrupt histone–DNA interactions, allowing transcriptional machinery and RNA polymerase II to bind and begin transcription.

Q: How does the SWI/SNF complex contribute to transcriptional activation?

SWI/SNF complexes are recruited to specific promoters by epigenetic marks on histones or by DNA-bound proteins. They disrupt histone–DNA interactions, opening the chromatin so that transcriptional machinery and polymerase II can access the DNA.

Q: Describe the model for transcriptional repression at a promoter.

A repressor recruits a corepressor and a histone deacetylase (HDAC) to remove acetyl groups, repressing transcription. Then a methyltransferase is recruited and methylates lysine 9 on histone H3 (K9 H3), leading to an inactive chromatin state and gene silencing.

Q: What is the role of HDACs in transcriptional repression?

HDACs (histone deacetylases) are recruited to specific gene loci by transcription factors. They remove acetyl groups from histones, causing chromatin condensation and gene silencing.

Q: What happens when K9 on histone H3 is methylated?

Methylation of lysine 9 on histone H3 leads to an inactive (closed) state of chromatin and gene silencing.

Q: How do Type II nuclear steroid hormone receptors differ from Type I?

Type II receptors (e.g., thyroid hormone receptor, retinoic acid receptor) are already located in the nucleus and bound to their DNA response elements even in the absence of ligand — unlike Type I receptors, which reside in the cytoplasm until ligand binds.

Q: How do Type II receptors regulate gene expression in the absence of ligand?

Without ligand, Type II receptors are bound to DNA and associated with NCoR and SMRT corepressor complexes and HDACs, which keep target genes repressed.

Q: What happens when ligand binds to a Type II nuclear receptor?

Ligand binding causes dissociation of the corepressors (NCoR/SMRT) and their replacement with coactivator complexes and histone acetyltransferases, which open chromatin and activate target genes.

Q: Why are steroid hormone signalling events important in the mammary gland microenvironment?

Steroid hormone signal transduction events are critical to reprogramming cells in the mammary gland, as the mammary gland microenvironment can reprogram cells of other tissues.