Question #1. Imprinting. Shown below is an imprinted gene complex called Snrpn I
ID: 64124 • Letter: Q
Question
Question #1. Imprinting. Shown below is an imprinted gene complex called Snrpn Imprinted Region. When the maternal ICR is imprinted (methylated), no lncRNA is made from Snrpn and the genes Ube3a and Atp10a are expressed. In contrast, when the ICR is unmethylated on the paternal chromosome, the Snrpn lncRNA is made and no Ube3a and Atp10a are made. However, four other genes (Frat3, Mkrn3, Magel2, and Ndn) are transcribed in a paternal specific pattern, hence these are paternal only expressed genes.
Answer the following questions regarding this imprinted region.
a. Indicated the state of methylation for the maternal and paternal ICR’s in each of the following tissues or cells at the particular developmental stages. Leave Open for unmethylated; Fill In for methylated. M=maternal; P=paternal
Inner Cell Mass of Embryo. Prior to implantation
M O O
P O O
PGC in genital ridge before sexual differentiation
M O O
P O O
Tissue of developing fetus
M O O
P O O
Spermatogonia, prior to meiosis in testis
M O O
P O O
Extraembryonic membrane of embryo
M O O
P O O
Zygote after nuclear fusion
M O O
P O O
b. Imprinting is responsible for directing expression of particular genes from one of the two chromosomes, maternal or paternal. As outlined, Ube3a and Atp10a are only expressed from the maternal chromosome. Ube3a codes for an ubitiquin ligase E6 protein that is only expressed in certain areas of the brain.
Describe a general molecular mechanism that would account for the tissue specific expression of Ube3a in the brain.
c. Consider the expression pattern of the Snrpn Complex and your understanding of imprinting. Indicate if the individual offspring would be affected, that is improper expression at imprint complex leading to a mutant phenotype. State Yes or No and give a reason.
1. Female offspring - Mutation in ICR that blocks it’s methylation – inherited from paternal chromosome.
Answer _____________________________________________________
Reason:
2. Male offspring - Deletion of the Ube3a and Atp10a genes – inherited from maternal chromosome.
Answer _______________________________________________________
Reason:
3. Male child - Deletion of the Frat3 to Ndn gene region – inherited from the maternal chromosome.
Answer __________________________________________________________
Reason:
4. Female child - Deletion of the ICR – inherited from the maternal chromosome
Answer _____________________________________________________________
Reason:
d. There is some evidence that imprinted genes can be altered in somatic tissue at a later point in the organism (following development). That is, the methylation imprint marks are erased and new patterns of gene expression are established in those cells. Outline the experimental approach you would use to analyze the methylation status of the ICR of the Snrpn imprinted gene region in the brain of mice at different periods after birth. Be brief.
e. Describe a molecular model (mechanism) for the cycle of imprinting at this imprinted region including:
How the imprint marks are removed and re established. Incorporate into your model possible ideas for how the Snrpn ICR would be protected from demethylation, then erased and then specifically re methylated. (Hint: names of enzymes must be included in your answer)
f. Suggest two possible models for how transcription of the lncRNA Snrnp leads to the paternal pattern of gene expression. With each model suggest an experimental approach that could be used to confirm the model.
Explanation / Answer
b.
Conceivable instruments for UBE3A impriting in the brain. (an) A guide of the maternal (MAT) and paternal (PAT) human chromosome locale 15q11-q13 containing UBE3A, adjusted from Lalande and Calciano [27]. Maternally expressed genes are portrayed in red and paternally expressed genes are delineated in blue. Non-engraved genes are represented to in green. Top: Methylation at the maternal PWS engraving focus (PWS-IC, dark circle) all around stifles articulation of surrounding genes (gray boxes), including the UBE3A antisense (UBE3A-ATS) transcript. On the other hand, the maternal copy of UBE3A is communicated (red arrow). Base: On the paternal chromosome, the PWS-IC contains a bunch of CpG destinations that are differentially methylated (open circle), allowing paternal genes expression (blue boxes), including the UBE3A-ATS transcript (blue bolt). The UBE3A-ATS (0.5-1.0 Mb long) covers the fatherly UBE3A locus, bringing about transcriptional hushing of UBE3A (red bolt blurring to white). Open triangles speak to the AS engraving focus (AS-IC). Neighboring genes upstream of UBE3A include: NDN (necdin) and qualities encoding snoRNAs [SNRPN, PAR5 (Prader-Willi Angelman Syndrome locale 5), HBII-8527, HBII-5247 and IPW (Imprinted in Prader-Willi syndrome)]. Neighboring qualities downstream of UBE3A include: ATP10A, the GABAA receptor b3, a5 and g2 subunits (GABRB3, GABRA5, GABRG3), OCA2 (Oculocutaneous albinism II) and HERC2 (Hect space and RDL 2). (b) Zoomed in area from (a) delineating a cutoff (dashed vertical blue line) past which the quieting of UBE3A translation by the UBE3A-ATS is fragmented [33]. Left of the line, the UBE3A-ATS transcript (dark blue shading) contends with the sense transcript (light red shading), bringing about quieting of full-length UBE3A sense transcripts. Conversely, to one side of the line, truncated paternal 5' portions of the UBE3A sense transcripts (red) are delivered [33]. (c) Hypothetical instruments of UBE3A-ATS/sense rivalry at the paternal allele. Top: Collision model [42]. On the off chance that translation can just happen in one course at a solitary time, RNA polymerases (RNAPII) deciphering the UBE3A sense strand (red) are contended off of their formats by approaching edifices occupied with interpretation of the UBE3A-ATS strand (blue). Base: RNA-DNA interaction model [42]. Generation of the UBE3A-ATS instigates histone adjustments (HM) that alter chromatin structural engineering along the UBE3A locus. Transcriptional extension of UBE3A is rashly prematurely ended at these locales, yielding truncated UBE3A sense transcripts (red).
f.
In eukaryotes, RNA transcription is a firmly directed procedure. NcRNAs can target diverse parts of this procedure, focusing on transcriptional activators or repressors, distinctive segments of the transcription reaction including RNA polymerase (RNAP) II and even the DNA duplex to manage gene interpretation and expression (Goodrich 2006). In mix these ncRNAs may contain an regulatory network that, including translation variables, finely control quality expression in complex eukaryotes.
NcRNAs balance the capacity of transcription variables by a few distinct mechanisms, including working themselves as co-controllers, altering interpretation element movement, or directing the affiliation and action of co-controllers. For instance, the ncRNA Evf-2 capacities as a co-activator for the homeobox interpretation variable Dlx2, which assumes imperative parts in forebrain advancement and neurogenesis (Feng 2006; Panganiban 2002)
Sonic hedgehog actuates transcription of Evf-2 from a ultra-monitored elementlocated between the Dlx5 and Dlx6 genes amid forebrain improvement (Feng 2006). Evf-2 then enrolls the Dlx2 transcription element to the same ultra-preserved component whereby Dlx2 along these lines affects articulation of Dlx5. The presence of other comparative ultra-or profoundly preserved components inside of the mammalian genome that are both translated and satisfy enhancer capacities propose Evf-2 may be illustrative of a summed up instrument that firmly manages essential formative genes with complex expression examples amid vertebrate development (Pennacchio 2006; Visel 2008). Without a doubt, the translation and articulation of comparative non-coding ultraconserved components was as of late appeared to be anomalous in human leukemia and to add to apoptosis in colon growth cells, proposing their association in tumorogenesis (Calin 2007).
Nearby ncRNAs can likewise select transcriptional programmes to manage adjoining protein-coding gene expression. The RNA tying protein TLS, ties and hinders the CREB tying protein and p300 histone acetyltransferease exercises on a curbed quality target, cyclin D1. The enrollment of TLS to the promoter of cyclin D1 is coordinated by long ncRNAs communicated at low levels and fastened to 5' regulatory regions in light of DNA harm signals (Wang 2008). Also, these nearby ncRNAs act helpfully as ligands to regulate the exercises of TLS. In the expansive sense, this component permits the cell to bridle RNA-tying proteins, which make up one of the biggest classes inside of the mammalian proteome, and incorporate their capacity in transcriptional programs. A late study found that a lncRNA in the antisense heading of the Apolipoprotein A1 (APOA1) manages the interpretation of APOA1 through epigenetic changes.
Late proof has raised the likelihood that interpretation of genes that escape from X-inactivation may be interceded by articulation of long non-coding RNA inside of the getting away chromosomal spaces