Transcription and RNA Processing Notes

RNA Types and Functions

• RNA Overview

  • Transcription is the process of creating RNA from DNA sequence, which plays a crucial role in the flow of genetic information within cells. The information is transcribed from the DNA sequence to produce various types of RNA essential for different cellular functions.

  • Types of RNA include coding (mRNA) and non-coding RNAs (ncRNA), each with specific roles in the gene expression pathway.

• Types of RNA

  • Messenger RNA (mRNA): Carries genetic code for protein synthesis, containing sequences that will be translated into amino acids. mRNA is synthesized from DNA during transcription and has a relatively short lifespan in the cell, typically ranging from minutes to hours, depending on the cellular context. mRNA is crucial for translating genetic information into functional proteins.

  • Ribosomal RNA (rRNA): Major component of ribosomes, which are the cellular machines that translate mRNA into proteins. rRNA makes up about 90% of cellular RNA, providing structural and functional foundations for ribosome function, playing a key role in facilitating peptide bond formation during translation.

  • Transfer RNA (tRNA): A small RNA molecule that transports specific amino acids to the ribosome during protein synthesis, ensuring that the genetic code is translated accurately. Each tRNA is specific to one amino acid and recognizes codons on the mRNA through its anticodon region.

  • Non-coding RNAs (ncRNA): Includes smaller types like microRNAs (miRNA) and small interfering RNAs (siRNA), which are involved in gene regulation and RNA processing. miRNAs regulate gene expression by targeting mRNA for degradation or inhibiting their translation, while siRNAs play a role in the RNA interference (RNAi) pathway, protecting cells from viral RNA and transposons.

  • Long non-coding RNAs (lncRNA): These are RNA molecules longer than 200 nucleotides that do not code for proteins but are involved in regulating gene expression at various levels, including chromatin remodeling, transcriptional control, and post-transcriptional processing.

Transcription Process

• Transcription Basics

  • RNA polymerase (RNAP) is the enzyme that synthesizes RNA from the DNA template, initiating the process of gene expression. RNAP unwinds the DNA double helix to access the template strand for transcription.

• Stages of Transcription

  • Initiation: RNAP binds to the promoter region of DNA, a specific DNA sequence that signals the start of a gene. This binding typically requires sigma factors in bacteria to facilitate the recognition of the promoter. The metabolic reconfiguration allows RNAP to create a transcription bubble where the DNA strands separate, providing access for the synthesis of the RNA strand.

  • Elongation: During elongation, RNAP synthesizes the RNA strand by adding nucleotides that are complementary to the template DNA strand. This synthesis occurs in the 5’ to 3’ direction, elongating the RNA molecule.

  • Termination: The end of RNA synthesis varies in mechanisms between prokaryotes and eukaryotes. In prokaryotes, termination can occur via intrinsic mechanisms (where RNA forms a hairpin loop) or Rho-dependent termination. In eukaryotes, termination usually involves polyadenylation, where a poly(A) tail is added to the 3’ end of mRNA, signaling the end of transcription.

• Role of Transcription Factors

  • General transcription factors (GTFs) assist in the process by recruiting RNA polymerase II (RNAP II) to the promoter, forming a pre-initiation complex (PIC).

  • TFII Class transcription factors, including TFIIA, TFIIB, and TFIID, contribute to the stabilization and regulation of the transcription machinery, ensuring proper transcription initiation. The Mediator Complex also plays a pivotal role in transducing signals from transcription factors to the RNAP II, integrating various regulatory inputs.

RNA Processing in Eukaryotes

• Key Post-Transcriptional Modifications

  • Capping: The addition of a 7-methylguanylate cap (m7G) to the 5’ end of the mRNA transcript protects it from degradation by exonucleases and facilitates nuclear export. The cap also plays a vital role in the initiation of translation by helping ribosomes recognize the mRNA.

  • Polyadenylation: The addition of a poly(A) tail to the 3’ end of the mRNA transcript enhances mRNA stability, influences nuclear export, and aids in translation. The length of the poly(A) tail can affect the lifespan of mRNA in the cytoplasm.

  • Splicing: Involves the removal of non-coding regions called introns and the joining of coding sequences known as exons. This process is facilitated by the spliceosome, a complex of snRNPs and other proteins that ensures precise splicing for mature mRNA.

• Splicing Mechanism:

  • Two transesterification reactions occur, resulting in a lariat structure as the introns are excised from the RNA transcript. The last step involves the ligation of exons, creating a continuous coding sequence that will be translated into a protein.

  • This process is highly regulated, and errors can lead to diseases, indicating its importance in cellular function.

Alternative Splicing

• Importance:

  • Alternative splicing allows a single gene to produce multiple protein isoforms, significantly increasing the diversity of the proteome without increasing the number of genes. This process enables cells to adapt to differing physiological conditions by changing the proteins they produce.

• Types:

  • Exon skipping: Removal of certain exons from the transcript, leading to the synthesis of proteins with different functional domains.

  • Alternative 3’ and 5’ splice site selection: The choice between different splice sites can create distinct mRNA variants from the same gene, influencing the resulting protein's function and properties.

  • Intron retention: May keep introns in the mature mRNA transcript, which can impact protein translation and functionality.

Conclusion

• Gene expression regulation relies heavily on the modulation of transcription and RNA processing, which are crucial for cellular homeostasis and response to environmental changes. Understanding these processes is critical for insights into cellular functions, developmental biology, and disease mechanisms, particularly in the context of various cancers and genetic disorders.