Gene Expression

Gene expression is the process by which information encoded in DNA is converted into functional products (primarily proteins). It involves transcription (DNA to mRNA), RNA processing, translation (mRNA to protein), and post-translational modification. Regulation occurs at every step and is critical for cell differentiation, development, and disease.

• Promoters: DNA sequences upstream of the transcription start site that recruit RNA polymerase II and general transcription factors. The TATA box (~25-30 bp upstream) is the classic core promoter element, though many human genes use TATA-less promoters with CpG islands.
• Enhancers and silencers: Cis-regulatory elements that increase or decrease transcription, respectively. Can function over large genomic distances (up to 1 Mb) and in either orientation. Enhancers bind tissue-specific transcription factors and physically interact with promoters through chromatin looping.
• RNA splicing: Removal of introns and joining of exons from pre-mRNA. Carried out by the spliceosome (snRNPs: U1, U2, U4, U5, U6). Splice sites are defined by consensus sequences: GT at the 5' (donor) splice site, AG at the 3' (acceptor) splice site, and a branch point adenosine. Alternative splicing allows one gene to produce multiple protein isoforms.
• 5' and 3' untranslated regions (UTRs): The 5' UTR contains regulatory elements affecting translation efficiency (e.g., upstream open reading frames, internal ribosome entry sites). The 3' UTR contains elements controlling mRNA stability, localization, and translation (e.g., AU-rich elements, miRNA binding sites, polyadenylation signals such as AAUAAA).
• mRNA processing: Three key modifications: (1) 5' cap (7-methylguanosine) added co-transcriptionally, which protects from degradation and facilitates translation initiation; (2) splicing removes introns; (3) 3' polyadenylation, in which cleavage and addition of ~200 adenine residues stabilizes mRNA and aids nuclear export.
• Nonsense-mediated decay (NMD): A quality control mechanism that degrades mRNAs containing premature termination codons (PTCs). Generally, a PTC is recognized if it is located >50-55 nucleotides upstream of the last exon-exon junction. NMD prevents translation of truncated, potentially toxic proteins. Clinically relevant: NMD-escaping variants may produce dominant-negative proteins (more severe phenotype), while NMD-triggering variants cause haploinsufficiency (may be milder).
• Post-translational modification (PTM): Chemical modifications that alter protein function, localization, or stability after translation. Key examples: phosphorylation (signal transduction), ubiquitination (targets for proteasomal degradation), glycosylation (protein folding, cell surface markers), acetylation (histone regulation), proteolytic cleavage (activation of zymogens like insulin from proinsulin).
• Gene regulation levels: Transcriptional (most common), post-transcriptional (splicing, mRNA stability, miRNA), translational (initiation factors, mTOR pathway), and post-translational (PTMs, protein degradation). Epigenetic regulation (DNA methylation, histone modification) provides an additional layer.

• Splice site variants: Pathogenic variants at GT/AG splice sites or within branch point sequences cause exon skipping, intron retention, or cryptic splice site activation. ~15-30% of all disease-causing mutations affect splicing. Example: some SMN1 variants in spinal muscular atrophy disrupt exon 7 splicing.
• Beta-thalassemia (HBB): Demonstrates mutations at virtually every level of gene expression, including promoter mutations (reduced transcription), splice site mutations (aberrant mRNA), nonsense mutations (premature termination/NMD), and frameshift mutations. The resulting severity correlates with residual beta-globin production (beta-plus vs beta-zero).
• Enhancer mutations: Disruption of limb-specific enhancers of SHH (ZRS enhancer) causes preaxial polydactyly. Deletion of an enhancer ~1 Mb from the gene demonstrates long-range regulatory control.
• NMD and phenotype prediction: In COL1A1, nonsense variants that trigger NMD cause haploinsufficiency and milder osteogenesis imperfecta (type I). Variants that escape NMD produce abnormal collagen chains with dominant-negative effects, causing more severe disease (types II-IV).