Ch 16 Gene Regulation in Eukaryotes 12 06 2014 12 06 2014 12 06 2014 17 Regulation of Gene Expression III Regulation of Gene Expression in Eukaryotes Eukaryotic genes are organized differently from those of prokaryotes The messages of eukaryotic genes are monocistronic Many genes contain intervening sequences introns that must be removed and exons that must be spliced together Processed mRNA contains a 5 G cap and 3 polyA tail Similar to prokaryotes there are some genes expressed constantly and some are highly regulated in their expression Eukaryotic genes are regulated by a set of regulatory DNA sequences recognized by trans acting factors Prokaryotes transcription and translation occur simultaneously for there is no nucleus to separate transcription from translation and no introns to be removed Eukaryotes gene expression can be regulated at the transcriptional level posttranscriptional level translational level or posttranslational level A Transcriptional regulation The rate of transcription is regulated at the gene level w different mechanism operating for long term and short term controls Long term transcriptional control o Chromosomes are condensed during cell division but gradually uncoil again to become dispersed as chromatin o A portion of chromatin remains condensed as heterochromatin not readily accessible by RNA polymerases and other proteins o Some regions are more diffuse existing as euchromatin and are accessible for transcription Barr bodies one of the X chromosomes in females is o Another way to control transcription on selected to chromosomes is inactivated to methylate the DNA Methylation is done by DNA methylases Deoxy cytosine methylase DCM or deoxy adenosine methylase DAM which add methyl groups to the C and A bases resp DNA methylation at the C and A nucleotides results in the prevention of such regions from being transcribed In humans as much as 5 of DNA is methylated and only about 3 5 of genes are actively transcribed at any given time DNA methylation also helps add DNA polymerase to distinguish the old strand from the new strand In bacteria DNA methylation is critical for recognizing cellular versus viral DNA Pattern and amount of methylation varies w species Short term transcriptional control o Some regions of chromosomes are actively transcribed as transcription puffs o Such regions can be easily seen in large multistranded polytene chromosomes in the salivary gland cells of Drosophila larva o Transcription of genes in eukaryotes is controlled by the interaction between transcription factors and the regulatory elements on the genes o The regulatory elements include the promoter which is closer to and upstream of the start site and enhancers or repressors which may be upstream or downstream from the start site o The promoter is present at a relatively fixed distance whereas other elements may be present close to or far from the start site o Trans acting proteins recognize specific cis acting elements on the DNA by binding to a specific DNA base sequence and inducing or inhibiting transcription o Some factors directly bind to RNA polymerase or other DNA binding proteins and affect transcription through protein protein interaction o Transcription factors have unique secondary structures to recognize the DNA Helix turn helix proteins contain alpha helix beta turn and alpha helix secondary structures that fit within the grooves of DNA protein sequences can vary and thus bind to different DNA sequences Zinc finger proteins these proteins contain zinc ions covalently bonded to certain residues which protrude as fingers and recognize DNA Leucine zippers contain multiple leucine residues which interact bc of the hydrophobic nature of their side chains o All these DNA binding proteins act as dimers which interact w each other along these motifs unique secondary tertiary structures and w DNA to regulate transcription o Some hormones can bind to these transcription factors in cytoplasm and then migrate to the nucleus bind w specific cis acting elements an activate the transcription of certain genes B Post transcriptional regulation Limited compared to transcriptional regulation Involves differential processing of the mRNA transcript or relative stabilities of mRNA Alternate splicing during mRNA processing introns are removed and exons are spliced o Some mRNAs are processed differently to yield different proteins mRNA stability some mRNAs have unique sequences at 5 or 4 NTRs that affect their stability in the cytoplasm o Some mRNAs are long lived several days some short few minutes o Long lived mRNAs allows translation to continue for a longer time o mRNA stability is altered by RNA binding proteins which may tag the mRNA for degradation or degrade the mRNA themselves C Translational regulation Translation in eukaryotes occurs in the cytoplasm and involves several proteins known as initiation factors and elongation factors mRNA can be translated more efficiently or less efficiently depending on the sequence of the ribosome recognition site on the mRNA Another translational control is to block translation of processed mRNA until the need arises mRNA storage many processed mRNAs are stored in the nucleus of an egg prior to fertilization and are translated after fertilization an during embryonic development o This control may be exerted by not modifying the 5 G cap or not having the right initiation factor present until the cells are ready to translate such mRNAs Hormonal regulation an alternate mechanism involves hormonal action Cofactor influence cofactors help increase the translational efficiency of certain mRNAs D Post translational modification Last stage of controlling gene expression at the protein level Results in functional proteins in most situations automatically Zymogen activation some proteins are first made in inactive forms and a In some situations it is used as a regulatory mechanism later they are converted to active forms Selective targeting for membrane proteins and proteins targeted for specific organelles to be active they should be in the right place o A segment of proteins normally at the NH2 terminal determines o The protein may then be transported through Golgi vesicles to the the protein s target site right membrane or organelle o Once it reaches its destination the target peptide also called the leader sequence or signal sequence is removed and the protein become active o Sometimes if the protein does not reach the target often bc of