Related Experiment Video

Updated: Jun 18, 2026

In Vitro Ubiquitination and Deubiquitination Assays of Nucleosomal Histones
11:36

In Vitro Ubiquitination and Deubiquitination Assays of Nucleosomal Histones

Published on: July 25, 2019

Ubiquitin ligase components Cullin4 and DDB1 are essential for DNA methylation in Neurospora crassa

Yuanbiao Zhao1, Ye Shen, Silu Yang

  • 1State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, China.

The Journal of Biological Chemistry
|December 2, 2009
PubMed

Insights

Ubiquitin ligase components Cullin4 and DDB1 are essential for DNA methylation in Neurospora. These proteins regulate DNA methylation by affecting histone H3K9 trimethylation, linking ubiquitin ligase activity to gene silencing mechanisms.

Area of Science:

  • Epigenetics
  • Molecular Biology
  • Fungal Genetics

Background:

  • DNA methylation and histone H3K9 trimethylation are key epigenetic marks involved in gene silencing and heterochromatin formation in eukaryotes.
  • In Neurospora crassa, histone methyltransferase DIM-5 catalyzes H3K9 trimethylation, which is essential for DNA methylation, with HP1 mediating the recruitment of DNA methyltransferase DIM-2.

Purpose of the Study:

  • To investigate the role of ubiquitin ligase components Cullin4 (Cul4) and DDB1 in DNA methylation and heterochromatin assembly in Neurospora.
  • To elucidate the mechanism by which Cul4 and DDB1 regulate H3K9 trimethylation and its connection to DNA methylation.

Main Methods:

  • Genetic analysis of Neurospora crassa mutants deficient in Cul4 and DDB1.
  • Western blotting and chromatin immunoprecipitation (ChIP) to assess H3K9 trimethylation levels.
  • Co-immunoprecipitation assays to determine in vivo interactions between Cul4, DDB1, and DIM-5.

Main Results:

  • Cullin4 and DDB1 are essential for DNA methylation in Neurospora.
  • The E3 ligase activity of the Cul4-based ubiquitin ligase is required for H3K9 trimethylation.
  • Cul4 and DDB1 physically associate with the histone methyltransferase DIM-5 in vivo.

Conclusions:

  • Cul4 and DDB1 regulate DNA methylation in Neurospora by influencing H3K9 trimethylation, likely through their association with DIM-5.
  • This study reveals a novel link between ubiquitin ligase machinery and the establishment of heterochromatin, suggesting a conserved regulatory mechanism across eukaryotes.

More Related Videos

Immunofluorescence Analysis of Endogenous and Exogenous Centromere-kinetochore Proteins05:35

Immunofluorescence Analysis of Endogenous and Exogenous Centromere-kinetochore Proteins

Published on: March 3, 2016

Immunostaining for DNA Modifications: Computational Analysis of Confocal Images09:42

Immunostaining for DNA Modifications: Computational Analysis of Confocal Images

Published on: September 7, 2017

Related Concept Videos

Covalently Linked Protein Regulators02:04

Covalently Linked Protein Regulators

Proteins can undergo many types of post-translational modifications, often in response to changes in their environment. These modifications play an important role in the function and stability of these proteins. Covalently linked molecules include functional groups, such as methyl, acetyl, and phosphate groups, and also small proteins, such as ubiquitin. There are around 200 different types of covalent regulators that have been identified.
These groups modify specific amino acids in a protein.
Nucleosome Remodeling02:54

Nucleosome Remodeling

Nucleosomes are the basic units of chromatin compaction. Each nucleosome consists of the DNA bound tightly around a histone core, which makes the DNA inaccessible to DNA binding proteins such as DNA polymerase and RNA polymerase. Hence, the fundamental problem is to ensure access to DNA when appropriate, despite the compact and protective chromatin structure.
Nucleosome remodeling complex
Eukaryotic cells have specialized enzymes called ATP-dependent nucleosome remodeling enzymes. These enzymes...
Anaphase Promoting Complex00:50

Anaphase Promoting Complex

The stepwise destruction of specific proteins is necessary for the progression and completion of the cell cycle. Such proteins are ubiquitinated by ubiquitin ligases and then subsequently destroyed by the proteasome. The SCF (Skp1/Cullin/F-box) and the anaphase-promoting complex (APC) are two important ubiquitin ligases involved in cell cycle progression. While SCF is active throughout the cell cycle, APC gets activated during metaphase to anaphase transition. Cdc20 or Cdh1 binds to APC and...
Nucleotide Excision Repair01:38

Nucleotide Excision Repair

DNA Distortion and Damage
Cells are regularly exposed to mutagens—factors in the environment that can damage DNA and generate mutations. UV radiation is one of the most common mutagens and is estimated to introduce a significant number of changes in DNA. These include bends or kinks in the structure, which can block DNA replication or transcription. If these errors are not fixed, the damage can cause mutations, which in turn can result in cancer or disease depending on which sequences are...
Nucleotide Excision Repair01:08

Nucleotide Excision Repair

Overview
Protein Complexes with Interchangeable Parts01:57

Protein Complexes with Interchangeable Parts

Groups of proteins may form a complex where each protein in this complex has a different role in the overall execution of the complex’s function. Often some of the proteins in the complex can be replaced by a closely related variant to give a complex that contains many of the same components yet is functionally distinct.
The SCF ubiquitin ligase is a protein complex of five individual proteins. This complex attaches ubiquitin to other target proteins to mark them for degradation. In order to...