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Benguet State University College of Natural Sciences DEPARTMENT OF BIOLOGY La Trinidad, Benguet Bio 50: Developmental...

Benguet State University College of Natural Sciences DEPARTMENT OF BIOLOGY La Trinidad, Benguet Bio 50: Developmental Biology Chapter 1. DEVELOPMENTAL GENETICS Lesson 3. Transcription Ribonucleic Acids (RNA) Messenger RNA (mRNA) Synthesis Eukaryotic RNA Processing LEARNING OBJECTIVES: By the end of this lesson, you should be able to: describe gene expression identify and explain the steps in the transcription process identify and explain the processes involved to form the eukaryotic mRNA The central dogma of molecular biology is a theory that states that – All cells, from bacteria to humans, the flow of genetic information in cells is from DNA to RNA to protein. Gene expression ▪ the process by which gene sequences are transcribed into functional gene products – proteins or functional RNAs (rRNA, tRNA, small RNA, etc.) ▪ two phases – transcription and translation Figure 1. An overview of the flow of information from DNA to protein in a eukaryote. CORTEZ, Sundoval E. Course Instructor Gene expression and regulation are the basis of cell development and differentiation. They also allow the cell to adapt to different conditions. By controlling the time, location, and expression level, gene transcripts can have a profound effect on the functions of genes within cells or in multicellular organisms. RIBONUCLEIC ACIDS (RNA) Comparison between RNA and DNA: RNA – ribose; DNA – deoxyribose T in DNA is replaced by U in RNA RNA – single-stranded and DNA – double-stranded (double helix); thus, RNA does not contain equal amounts of specific bases RNA is much smaller (75 to < thousand nucleotides) than DNA Figure 2. Differences and similarities in DNA and RNA TYPES of RNA Five major types, distinguished by their function: 1. Heterogeneous nuclear RNA (hnRNA) ▪ RNA formed directly by DNA transcription; RNA processing converts hnRNA to mRNA 2. Messenger RNA (mRNA) ▪ RNA that carries instructions for protein synthesis (genetic information) to the sites for protein synthesis 3. Small nuclear RNA (snRNA) ▪ RNA that facilitates the conversion of hnRNA to mRNA 4. Ribosomal RNA (rRNA) ▪ RNA that combines with specific proteins to form ribosomes, the physical sites for protein synthesis ▪ rRNA present in ribosomes has no informational value 5. Transfer RNA (tRNA) ▪ RNA that delivers amino acids to the sites for protein synthesis CORTEZ, Sundoval E. Course Instructor MESSENGER RNA (mRNA) SYNTHESIS Transcription ▪ the process by which DNA directs the synthesis of hnRNA/mRNA molecules that carry the coded information needed for protein synthesis ▪ occurs in the nucleus Gene ▪ a segment of a DNA strand that contains the base sequences for the production of a specific hnRNA/mRNA molecule STEPS in the TRANSCRIPTION PROCESS 1. Initiation A portion of the DNA double helix unwinds, exposing some bases (a gene). The unwinding process is governed by the enzyme RNA polymerase. The DNA sequence onto which the proteins and enzymes involved in transcription bind to initiate the process is called a promoter. In most cases, promoters exist upstream of the genes they regulate. Figure 3. The initiation of transcription begins when DNA is unwound, forming a transcription bubble. Enzymes and other proteins involved in transcription bind at the promoter. Transcription bubble ▪ the region of unwinding or opened-up DNA RNA polymerase ▪ enzyme that performs transcription ▪ governs the unwinding process ▪ catalyzes formation of phosphodiester bonds that links ribonucleotides together to form a linear chain ▪ extend the growing RNA chain by one nucleotide at a time in the 5’ to 3’ direction 2. Elongation Free ribonucleotides, one nucleotide at a time, align along one of the exposed strands of DNA bases, the template strand, forming new base pairs. In this process, U rather T aligns with A in the base- pairing process. RNA polymerase is involved in the linkage of ribonucleotides, one by one, to the growing hnRNA molecule. Figure 4. During elongation, RNA polymerase tracks along the DNA template, synthesizes mRNA in the 5' to 3' direction, and unwinds then rewinds the DNA as it is read. CORTEZ, Sundoval E. Course Instructor  3. Termination Transcription ends when the RNA polymerase enzyme encounters a sequence of bases that is “read” as a stop signal. The newly formed hnRNA molecule and the RNA polymerase enzyme are released, and the DNA then rewinds to re-form the original double helix. Figure 5. Processes for synthesis of functional mRNA in prokaryotes and eukaryotes. EUKARYOTIC RNA PROCESSING RNA processing ▪ the sequence of events through which the primary transcript from a gene acquires its mature form 1. Covalent modification of the ends of the RNA Both ends of eukaryotic mRNAs are modified by Capping on the 5’ end ▪ first modification of eukaryotic pre-mRNAs ▪ the cap consists of a modified guanine nucleotide Polyadenylation of the 3’ end An enzyme called poly-A polymerase adds a chain of adenine nucleotides to the RNA. Figure 6. Eukaryotic mRNA contains introns that must be spliced out. A 5' cap and 3' tail are also added. CORTEZ, Sundoval E. Course Instructor  2. Removal of intron sequences discarded from the middle of the RNA transcript by the process of RNA splicing A gene is segmented consisting of alternating exons and introns. Thus, not all bases in a gene convey genetic information. Exon ▪ a gene segment that conveys (codes for) genetic information; DNA segments that help express a genetic message Intron ▪ a gene segment that does not convey (code for) genetic information; DNA segment that interrupts a genetic message Both the exons and the introns of a gene are transcribed during production of hnRNA. Figure 7. Exons and introns RNA splicing ▪ functions in the production of mRNA ▪ introns are removed from the newly synthesized RNA ▪ joins together the exons RNA splicing involves snRNA complexed with proteins in particles called small nuclear ribonucleoprotein particles (snRNP). It further collects together into larger complexes called spliceosomes that now converts hnRNA molecules to mRNA molecules. CORTEZ, Sundoval E. Course Instructor Figure 8. An overview of alternative splicing. This process can produce a diversity of mRNAs from a single gene by arranging coding sequences (exons) from recently spliced RNA transcripts into different combinations. How does the cell distinguish between the mRNA molecules to keep and the potentially dangerous debris (excised introns, broken RNAs, and aberrantly processed pre-mRNAs) from RNA processing? Improperly processed mRNAs, and other RNA debris are retained in the nucleus, where they are eventually degraded by the nuclear exosome, a large protein complex whose interior is rich in exonucleases. Eukaryotic cells thus export only useful RNA molecules to the cytoplasm, while debris is disposed of in the nucleus Nuclear pore complexes (NPCs) ▪ aqueous channels in the nuclear membrane that directly connect the nucleoplasm and cytosol where mRNAs are guided through. Figure 9. The nucleus CORTEZ, Sundoval E. Course Instructor