Human DROSHA Structure & MicroRNA Biogenesis | RNA Scissors

RNA secondary structure forms when complementary base pairs along a single strand create double-helix regions called stems, while unpaired bases form loops. Common structural elements include: (1) Bulge loops - regions where one strand extends beyond the other without pairing; (2) Internal loops - gaps where bases cannot find complementary partners; (3) Multibranch junctions - complex branching points where multiple RNA strands interact; (4) Stem-loop structures - the most common form where a double-stranded stem connects to a single-stranded loop. These punctuated structures arise because RNA must fold to protect vulnerable bases from damage and enable functional interactions.

Stem-loop (hairpin) structures form when single-stranded nucleic acids fold back on themselves, creating paired double-helix regions (stem) and unpaired loops. Formation requires palindromic sequences where the 5' end base pairs with the corresponding 3' base. GC base pairs (three hydrogen bonds) form more stable stems than AU pairs (two hydrogen bonds). To identify sequences capable of forming stable stem-loops, check whether complementary bases align at both ends of the sequence. Sequences with higher GC content and better palindromic symmetry will form more stable hairpin structures, which are common in RNA secondary structures including tRNA, rRNA, and viral RNAs.

This section covers RNA secondary structures and their biological significance. RNA forms various secondary structures through folding of its primary structure, including stem-loop (hairpin) structures, multi-branch junctions, internal loops, bulges, and pseudoknots. The stem-loop structure is the most common RNA secondary structure, formed when complementary sequences fold back on themselves to create a double-stranded stem connected to a single-stranded loop. These structures are essential for RNA function and play important roles in gene expression regulation and cellular processes. RNA secondary structures differ from DNA in that RNA contains ribose sugar (with C3' endo conformation) and can form more diverse structures due to its single-stranded nature and ability to fold into complex three-dimensional shapes.

RNA molecules adopt several characteristic secondary structural motifs including bulge loops, internal loops, multibranch junctions, and stem-loops. Bulge loops form when one strand folds back while leaving certain bases unpaired due to incorrect orientation. Internal loops occur when regions contain bases unable to find complementary partners, creating gaps in the paired regions. Multibranch junctions resemble four-way crossings where multiple strands interact. Stem-loops consist of paired stems (from complementary base pairing) connected by unpaired loops. All these structures share the common principle of alternating paired and unpaired regions throughout the RNA molecule.

Unlike DNA which exists as a double helix, RNA molecules predominantly exist as single-stranded molecules that fold into characteristic three-dimensional structures called stem-loop forms; these structures consist of complementary nucleotide sequences forming hydrogen-bonded stems and non-complementary sequences creating destabilized loops, enabling RNA to form secondary and tertiary structures that allow it to function as biological catalysts, such as in ribosomes during protein synthesis.

RNA interference (RNAi) is a post-transcriptional gene regulation mechanism where small RNA molecules (miRNA, siRNA, shRNA) guide the RNA-induced silencing complex (RISC) to target specific mRNA sequences, leading to either mRNA cleavage or translational blockage, thereby preventing the synthesis of specific proteins; this process is crucial for defense against viral infections, regulation of gene expression, and maintenance of genomic stability.

RNA interference (RNAi) is a powerful cellular mechanism used by many organisms to control gene expression by preventing protein production. Two key types of regulatory RNAs are involved: small interfering RNAs (siRNAs) and microRNAs (miRNAs). The process begins when genes are transcribed from DNA to RNA in the nucleus, then processed and transported to the cytoplasm as messenger RNA (mRNA), which is translated into proteins by ribosomes. In the cytoplasm, siRNAs and miRNAs exert suppressive effects. Both types originate from longer double-stranded RNAs, bind to the Argonaute enzyme, and are cut into ~21-nucleotide fragments. These fragments form the RNA-induced silencing complex (RISC), which targets specific mRNAs. siRNAs bind perfectly complementarily to target mRNAs, causing cleavage and degradation. miRNAs bind via seed regions, allowing them to regulate hundreds of different RNAs. RNAi prevents protein formation and plays crucial roles in plants, animals, fungi, and bacteria, with promising applications in molecular biology and medicine.

RNA interference (RNAi) is a fundamental cellular mechanism used by organisms to control gene expression with high precision. The process involves small RNA molecules that guide gene silencing complexes to specific messenger RNAs. Two main types of regulatory RNAs exist: small interfering RNAs (siRNAs), derived from double-stranded RNAs, and microRNAs (miRNAs), transcribed from nuclear precursors. Both types are processed by Dicer into ~21-nucleotide fragments that bind Argonaute proteins, forming the RNA-induced silencing complex (RISC). siRNAs typically have perfect complementarity and cleave target mRNAs, while miRNAs use partial matching (seed region) to target hundreds of mRNAs, often inhibiting translation rather than causing degradation. This mechanism is evolutionarily conserved across plants, animals, fungi, and some bacteria, playing crucial roles in development, immunity, and disease prevention, while also serving as powerful tools for laboratory research and potential therapeutic applications.

RNA interference is a mechanism where microRNA regulates gene expression by binding to mRNA and preventing its translation into protein. This process is crucial for controlling gene expression and maintaining cellular homeostasis. The discovery of RNA interference and microRNA was awarded the Nobel Prize in 2024, highlighting its importance in understanding gene regulation. Understanding RNA interference is important for understanding gene regulation and has implications for developing new therapeutic approaches.

RNA interference (RNAi) is a mechanism by which small RNA molecules (miRNA and siRNA) regulate gene expression by binding to complementary mRNA sequences and causing their degradation or translational repression. The instructor emphasizes that understanding RNAi is essential for understanding how genes are regulated and how cells control protein production. This mechanism is crucial for cellular development and response to environmental stimuli.

Endonucleases are enzymes that cleave DNA internal to its sequence, operating on both linear and circular DNAs. They encompass restriction endonucleases (classified into Types I-IV), homing endonucleases, DNases, and RNases. Type I enzymes protect against foreign DNA through poorly defined patterns. Type II enzymes (most commonly used) are sequence-specific, generating diverse overhangs including four-base pairs (BamHI, XbaI, EcoRI), two-base pairs, blunt ends, or variable lengths. Type III and IV enzymes have specialized functions. Advanced technologies like Type II-S (BsaI) and Type II-G (Bc1) enable scarless ligation by removing restriction sites entirely. Practical considerations include star activity, buffer specificity, supercoiled DNA requirements, and methylation effects. All restriction enzymes generate consistent chemistry: three prime hydroxy and five prime phosphate ends. Homing endonucleases provide precise genome editing tools. DNAse I enables DNA removal from RNA samples and techniques like nick translation. RNases serve specialized roles: RNAse A removes RNA contamination, RNAse H degrades RNA-DNA hybrids, and RNAse E processes mRNAs and tRNAs.

DNA cleavage enzymes are categorized into three types based on their cutting mechanism. Exonucleases cut DNA from the ends (5' to 3' direction), removing nucleotides one at a time. Endonucleases cut within the DNA helix, breaking the double helix structure between nucleotides. Restriction endonucleases recognize specific palindromic sequences and cleave DNA at these sites, making them essential tools in recombinant DNA technology for precise DNA fragmentation.

Nucleases are enzymes that cut nucleic acids (DNA and RNA) and are classified into two main types: (1) Exonucleases - cut nucleic acids from the ends (termini); (2) Endonucleases - cut nucleic acids at internal sites within the molecule. Exonucleases remove nucleotides one by one from the ends, while endonucleases make cuts within the DNA/RNA chain. Endonucleases are further classified into normal endonucleases and restriction endonucleases. Restriction endonucleases are also called 'molecular scissors' because they cut DNA at specific recognition sequences.

Nucleic acid-cleaving enzymes are categorized into two main types: exonucleases and endonucleases. Exonucleases remove nucleotides from the ends of DNA or RNA molecules and are classified into three types (Exonucleases 1, 2, and 3), each with distinct mechanisms of action. Endonucleases cleave nucleic acids at internal sites within the molecule, with restriction endonucleases being a major subclass. Restriction endonucleases are further divided into three types: Type 1 (requiring ATP and magnesium ions, cutting at variable distances from recognition sites), Type 2 (requiring only magnesium ions, cutting at specific sites within recognition sequences, most commonly used in molecular biology), and Type 3 (requiring ATP and magnesium ions, cutting at variable distances from recognition sites). The key difference between exonucleases and endonucleases lies in their cleavage location: exonucleases act on terminal ends while endonucleases act on internal sites.

RNase III enzymes Drosha and Dicer, traditionally known for microRNA biogenesis, have additional functions independent of microRNAs: Drosha cleaves messenger RNAs containing stem-loop structures to regulate gene expression (demonstrated in neuronal progenitor maintenance via neurogenin2 repression), while Dicer degrades cytotoxic retrotransposon-derived RNAs (Alu/B1/B2 elements) to protect retinal epithelial cells from damage in diseases like geographic atrophy.