Related Experiment Video

Updated: Aug 8, 2026

Mutagenesis and Analysis of Genetic Mutations in the GC-rich KISS1 Receptor Sequence Identified in Humans with Reproductive Disorders
12:49

Mutagenesis and Analysis of Genetic Mutations in the GC-rich KISS1 Receptor Sequence Identified in Humans with Reproductive Disorders

Published on: September 4, 2011

Forskolin-resistant Y1 mutants harbor defects associated with the guanyl nucleotide-binding regulatory protein, Gs

B P Schimmer1, J Tsao, R Borenstein

  • 1Banting and Best Department of Medical Research, University of Toronto, Ontario, Canada.

The Journal of Biological Chemistry
|November 15, 1987
PubMed

Insights

Forskolin-resistant Y1 cell mutants show decreased adenylate cyclase activity and altered responses to hormones, indicating a defect in the Gs protein pathway. This research helps pinpoint the mutation site affecting cellular signaling.

Area of Science:

  • Cellular Biology
  • Molecular Endocrinology
  • Signal Transduction

Background:

  • Adenylate cyclase activity is crucial for cellular responses.
  • Forskolin is a known activator of adenylate cyclase.
  • Y1 adrenocortical tumor cells are a model for studying steroidogenesis.

Purpose of the Study:

  • To compare adenylate cyclase properties in forskolin-resistant Y1 cell mutants and parental Y1 cells.
  • To identify the molecular site of mutation responsible for forskolin resistance.
  • To elucidate the role of Gs protein in forskolin resistance.

Main Methods:

  • Enzyme kinetics analysis of adenylate cyclase.
  • Assessment of enzyme response to forskolin, NaF, and guanyl-5'-yl imidodiphosphate (Gpp(NH)p).
  • Cholera toxin and pertussis toxin-catalyzed ADP-ribosylation assays.

Main Results:

  • Mutant adenylate cyclase exhibited reduced activity and altered kinetics compared to parental cells.
  • Mutant enzymes were insensitive to ACTH and less responsive to Gpp(NH)p.
  • Mutant membranes showed significantly reduced cholera toxin-catalyzed ADP-ribosylation, while pertussis toxin labeling remained unchanged.

Conclusions:

  • The mutation confers resistance to forskolin by affecting adenylate cyclase activity.
  • A defect in the Gs protein is implicated in the forskolin resistance phenotype.
  • These findings localize the mutation to a component of the Gs-adenylate cyclase signaling pathway.

More Related Videos

Comparing the Affinity of GTPase-binding Proteins using Competition Assays10:37

Comparing the Affinity of GTPase-binding Proteins using Competition Assays

Published on: October 8, 2015

Deciphering the Structural Effects of Activating EGFR Somatic Mutations with Molecular Dynamics Simulation15:05

Deciphering the Structural Effects of Activating EGFR Somatic Mutations with Molecular Dynamics Simulation

Published on: May 20, 2020

Related Concept Videos

GTPases and their Regulation02:14

GTPases and their Regulation

Guanine nucleotide-binding proteins (G-proteins), also known as GTPases, are a superfamily of proteins that regulate many cellular processes, such as cell signaling, vesicular transport, and the regulation of cell shape and motility. Mutation or dysfunction of these proteins can lead to disease. There are around 40,000 known G-proteins that can broadly be classified into two groups ‒  small G-proteins consisting of a single domain and large multi-domain G-proteins.
Large G-proteins, also known...
The Ras Gene02:38

The Ras Gene

The Ras-gene-encoded proteins are regulators of signaling pathways controlling cell proliferation, differentiation, or cell survival. The Ras-gene family in humans constitutes three primary members—the HRas, NRas, and KRas. These genes code for four functionally distinct yet closely related proteins—the HRas, NRas, KRas4A, and KRas4B. The involvement of mutant Ras genes in human cancer was first discovered in 1982 and is among the most common causes of human tumorigenesis.
Ras is a superfamily...
Small GTPases - Ras and Rho01:24

Small GTPases - Ras and Rho

Ras and Rho are small monomeric GTPases that act downstream of receptor tyrosine kinase (RTK) and regulate various cellular processes. These GTPases switch between active and inactive states by binding to guanine nucleotides.
Three regulatory proteins control their activity:
Activation and Inactivation of G Proteins01:22

Activation and Inactivation of G Proteins

Heterotrimeric G proteins are guanine nucleotide-binding proteins. As the name suggests, heterotrimeric G proteins are composed of three subunits: alpha, beta, and gamma. They remain GDP-bound or GTP-bound inside the cells and switch between inactive/active states. The Gα subunit possesses the nucleotide-binding pocket that binds guanine nucleotides and switches between GDP or GTP-bound states. In contrast, the Gꞵ and Gγ subunits are always bound together with high affinity and are together...
GPCR Desensitization01:12

GPCR Desensitization

G protein-coupled receptor (GPCR) signaling plays a crucial role in cell functioning. GPCR desensitization is an equally essential process. It allows cells to respond to changing environments and regain sensitivity to new stimuli while preventing unnecessary stimulation when no longer needed. Prolonged exposure to stimuli leads to GPCR desensitization. It involves blocking the receptors from binding and activating additional G proteins. This inhibits activation of downstream effectors, thereby...
GPCRs Regulate Adenylyl Cylase Activity01:09

GPCRs Regulate Adenylyl Cylase Activity

Some GPCRs transmit signals through adenylyl cyclase (AC), a transmembrane enzyme. AC helps synthesize second messenger cyclic adenosine monophosphate (cAMP). AC catalyzes cyclization reaction and converts ATP to cAMP by releasing a pyrophosphate. The pyrophosphate is further hydrolyzed to phosphate by the enzyme pyrophosphatase, which drives cAMP synthesis to completion. However, cAMP is rapidly degraded to 5′ AMP by the enzymes phosphodiesterase (PDE), preventing overstimulation of cells.
Two...