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Cis-regulatory Sequences

Cis-regulatory Sequences

Cis-regulatory sequences are short fragments of non-coding DNA that are present on the same chromosomes as the genes that they regulate. These fragments serve as binding sites for transcriptional regulators, proteins that are responsible for controlling gene transcription and differential gene expression across cell types in eukaryotes. Cis-regulatory sequences can be close to the gene of interest or thousands of bases away in the DNA sequence; however, those sequences that are further away are...

Cis-regulatory Sequences

Cis-regulatory Sequences

Cis-regulatory sequences are short fragments of non-coding DNA that are present on the same chromosomes as the genes that they regulate. These fragments serve as binding sites for transcriptional regulators, proteins that are responsible for controlling gene transcription and differential gene expression across cell types in eukaryotes. Cis-regulatory sequences can be close to the gene of interest or thousands of bases away in the DNA sequence; however, those sequences that are further away are...

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The organization of prokaryotic genes in their genome is notably different from that of eukaryotes. Prokaryotic genes are organized, such that the genes for proteins involved in the same biochemical process or function are located together in groups. This group of genes, along with their regulatory elements, are collectively known as an operon. The functional genes in an operon are transcribed together to give a single strand of mRNA known as polycistronic mRNA.
Transcription of prokaryotic...

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Applying an Inducible Expression System to Study Interference of Bacterial Virulence Factors with Intracellular Signaling

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Cryptic transcripts from a ubiquitous plasmid origin of replication confound tests for cis-regulatory function.

Nathan A Lemp¹, Kei Hiraoka, Noriyuki Kasahara

¹Department of Molecular and Medical Pharmacology, University of California, Los Angeles, CA 90095, USA.

Nucleic Acids Research

|May 24, 2012

Summary

Plasmid reporter assays can yield false positives due to cryptic promoters producing RNA. This readthrough RNA, when spliced, mimics gene regulation elements, potentially misattributing function.

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Quantitative Comparison of cis-Regulatory Element (CRE) Activities in Transgenic Drosophila melanogaster

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Published on: December 19, 2011

Area of Science:

Molecular Biology
Gene Regulation Studies

Background:

Plasmid reporter assays are crucial for studying gene expression regulation.
Previous studies have relied heavily on these assays to understand regulatory mechanisms.

Purpose of the Study:

To investigate a potential artifact in widely used plasmid reporter assays.
To identify the source of spurious reporter gene activation.
To re-evaluate the function of certain human sequences previously identified as regulatory elements.

Main Methods:

Analysis of RNA production from cryptic promoters within replication origins of common plasmids.
Assessment of reporter gene activity influenced by readthrough transcripts.
Utilizing dicistronic and monocistronic reporter assays (luciferase).
Investigating the role of 3' splice sites (ss) in readthrough transcript processing.
Computational estimation of cryptic 3' ss frequency in genomic sequences.

Main Results:

Widely used plasmids constitutively produce substantial RNA from a cryptic promoter.
Readthrough transcripts significantly stimulate reporter activity when containing a 3' splice site (ss).
Two human sequences, previously identified as regulatory elements, function by enabling splicing of these cryptic transcripts.
This mechanism creates a false appearance of promoter activity in reporter assays.
A strong correlation exists between reporter activity and spliced readthrough transcript levels.
Cryptic 3' ss are frequent in genomic sequences, suggesting widespread potential for misattribution.

Conclusions:

Common plasmid reporter assays can generate artifacts due to cryptic promoter activity and splicing.
The mechanism involves readthrough transcripts from cryptic promoters being spliced, mimicking regulatory elements.
This can lead to misattribution of cis-regulatory function to sequences containing 3' splice sites.
Caution is advised when interpreting results from such reporter assays; re-evaluation may be necessary.