Coexpression of mutant and wild type protein kinase in lymphoma cells resistant to dibutyryl cyclic AMP

Related Experiment Video

Updated: Aug 13, 2026

Assaying the Kinase Activity of LRRK2 in vitro
06:09

Assaying the Kinase Activity of LRRK2 in vitro

Published on: January 18, 2012

I Lemaire, P Coffino

    Journal of Cellular Physiology
    |September 1, 1977
    PubMed

    Insights

    A mutant mouse lymphoma cell line resistant to dibutyryl cyclic AMP was identified. This resistance stems from a mutated regulatory subunit of cyclic AMP-dependent protein kinase, affecting enzyme activity.

    Area of Science:

    • Cell Biology
    • Molecular Genetics
    • Biochemistry

    Background:

    • S49 mouse lymphoma cells are a model system for studying cyclic AMP-dependent pathways.
    • Cyclic AMP-dependent protein kinase (PKA) plays a crucial role in cellular signaling.
    • Understanding PKA regulation is vital for deciphering cellular responses to various stimuli.

    Purpose of the Study:

    • To isolate and characterize a mutant S49 cell line resistant to dibutyryl cyclic AMP.
    • To investigate the molecular basis of this resistance in cyclic AMP-dependent protein kinase.
    • To elucidate the genetic and functional properties of PKA regulatory subunits.

    Main Methods:

    • Isolation of dibutyryl cyclic AMP-resistant mutants from S49 cells.
    • Biochemical assays to assess cyclic AMP-dependent protein kinase activity and activation kinetics.
    • Cell hybridization experiments to analyze complementation of mutant and wild-type phenotypes.
    • Thermal stability assays to compare mutant and wild-type kinase properties.

    Main Results:

    • A mutant clone resistant to dibutyryl cyclic AMP was successfully isolated.
    • The mutant kinase exhibited reduced sensitivity to cyclic AMP activation and increased thermal lability compared to wild type.
    • Cell hybrids demonstrated resistance and expressed both mutant and wild-type kinase activities.
    • Parent mutant cells showed residual wild-type kinase activity.

    Conclusions:

    • The mutation affects the regulatory subunit of cyclic AMP-dependent protein kinase.
    • Wild-type S49 cells possess two identical alleles for the PKA regulatory subunit.
    • One allele is mutated in the resistant clone, leading to altered kinase function and cellular resistance.

    More Related Videos

    Methods for Evaluating the Role of c-Fos and Dusp1 in Oncogene Dependence10:09

    Methods for Evaluating the Role of c-Fos and Dusp1 in Oncogene Dependence

    Published on: January 7, 2019

    Characterize Disease-related Mutants of RAF Family Kinases by Using a Set of Practical and Feasible Methods07:49

    Characterize Disease-related Mutants of RAF Family Kinases by Using a Set of Practical and Feasible Methods

    Published on: July 17, 2019

    Related Concept Videos

    Inhibition of Cdk Activity02:34

    Inhibition of Cdk Activity

    The orderly progression of the cell cycle depends on the activation of Cdk protein by binding to its cyclin partner. However, the cell cycle must be restricted when undergoing abnormal changes. Most cancers correlate to the deregulated cell cycle, and since Cdks are a central component of the cell cycle, Cdk inhibitors are extensively studied to develop anticancer agents. For instance, cyclin D associates with several Cdks, such as Cdk 4/6, to form an active complex. The cyclin D-Cdk4/6 complex...
    Abnormal Proliferation02:23

    Abnormal Proliferation

    Under normal conditions, most adult cells remain in a non-proliferative state unless stimulated by internal or external factors to replace lost cells. Abnormal cell proliferation is a condition in which the cell's growth exceeds and is uncoordinated with normal cells. In such situations, cell division persists in the same excessive manner even after cessation of the stimuli, leading to persistent tumors. The tumor arises from the damaged cells that replicate to pass the damage to the daughter...
    mTOR Signaling and Cancer Progression03:03

    mTOR Signaling and Cancer Progression

    The mammalian target of rapamycin or mTOR protein was discovered in 1994 due to its direct interaction with rapamycin. The protein gets its name from a yeast homolog called TOR. The mTOR protein complex in mammalian cells plays a major role in balancing anabolic processes such as the synthesis of proteins, lipids, and nucleotides and catabolic processes, such as autophagy in response to environmental cues, such as availability of nutrients and growth factors.
    The mTOR pathway or the...
    Interactions Between Signaling Pathways01:19

    Interactions Between Signaling Pathways

    Signaling cascades usually lack linearity. Multiple pathways interact and regulate one another, allowing cells to integrate and respond to diverse environmental stimuli.
    Convergence and divergence, and cross-talk between signaling pathways
    Two distinct signaling pathways can converge on a single functional unit, which may either be a single protein or a complex of proteins. The response is either functionally distinct or synergistic between the two pathways but different from the response...
    Amplifying Signals via Enzymatic Cascade01:22

    Amplifying Signals via Enzymatic Cascade

    When a ligand binds to a cell-surface receptor, the receptor's intracellular domain changes shape, which may either activate its enzyme function or allow its binding to other molecules. The initial signal is amplified by most signal transduction pathways. This means that a single ligand molecule can activate multiple molecules of a downstream target. Proteins that relay a signal are most commonly phosphorylated at one or more sites, activating or inactivating the protein. Kinases catalyze the...
    PI3K/mTOR/AKT Signaling Pathway01:22

    PI3K/mTOR/AKT Signaling Pathway

    The mammalian target of rapamycin  (mTOR) is a serine/threonine kinase that regulates growth, proliferation, and cell survival in response to hormones, growth factors, or nutrient availability. This kinase exists in two structurally and functionally distinct forms: mTOR complex 1  (mTORC1) and mTOR complex 2  (mTORC2). The first form (mTORC1) is composed of a rapamycin-sensitive Raptor and proline-rich Akt substrate, PRAS40. In contrast,  mTORC2 consists of a rapamycin-insensitive companion...