Essay On Extracellular Trafficking

The goal of this project is to determine the effect of the RNAi targeting the gene unc-22 in C. Elegans through visual observation of worm

MicroRNAs (miRNA) are small noncoding RNA, usually 17-25 nucleotides long that are able to bind complementary sequences of target messenger RNA (mRNA) and to induce both their degradation and translational repression (Fortunato, et al 2014). They are one of the most significant classes of non-coding RNA molecules (eg. small interfering RNA (siRNA) and ribozymes) that act within the cell. MiRNAs are also evolutionary conserved in different species from plants to humans and are encoded by their respective genes (Bader, 2012).

1a. To select sRNA, mRNA, and lncRNA unique to Cn but not present in mammalian hosts.

Cardiovascular disease (CVD) is the major macrovascular complication of diabetes mellitus as well as the main cause of death from clinical observation. There are about 75% of patients with type 2 diabetes have died of myocardial infarction as reported. Non-coding RNA (ncRNA) is an RNA molecule that is not translated into a protein, which include snoRNAs, microRNAs, siRNAs, piRNAs, lncRNAs and others. Recently, increasing number of evidences implicate that these ncRNAs are crucially involved in many cellular responses as well as physiological and pathological processes of several diseases. In this review, we summarize recent progress in research on functional regulatory of series of ncRNAs in cardiovascular disease with Diabetes Mellitus. Meanwhile,

Ontology analysis and KEGG function annotations of the host target genes suggested that the miRNAs are involved in

Post-transcriptional gene regulation by sRNAs may occurs in various patterns of base-pairing with a target RNA resulting in different outcomes or by directly binding to proteins to modulate their function.12-14 Two classes of sRNAs are identified: trans-encoded RNAs which are transcribed from intergenic regions of the genome, and cis-encoded RNAs which are encoded on the strand complementary to coding sequences or the 5′ or 3′ untranslated region (5′ UTR, 3′ UTR) of transcripts.15-17 The family of trans-encoded sRNAs usually requires the chaperone Hfq to stabilize the often imperfect base-pairing interaction with target mRNA.18 In contrast, cis-encoded sRNAs possess a region of perfect complementarity to their target mRNA and Hfq is not needed for target binding. It shows that sRNAs are involved in many important physiological processes including anaerobic growth, nutrient availability, iron homeostasis and the response to oxidative, envelope and osmotic

However, the mechanism regulating the migration of mature ecRNAs into the cytoplasm remains unclear. The linear counterparts of circRNAs, such as mRNAs or lncRNAs, can penetrate the nuclear membrane through the nuclear pore complex. Therefore, some researchers put forward the hypothesis that ecRNA export may be regulated by a mechanism similar to the regulatory mechanism of linear RNA migration [18]. The degradation pathway of circRNAs is another unanswered question. The expression level of circRNA is dynamically modulated by the balance between biogenesis and degradation of circRNAs. CircRNAs may be degraded by short interfering RNAs [13]. A recent study found that circRNAs may be cleared by extracellular vesicles or microvesicle release in mammalian cells [43]. However, this conclusion is based on an in vitro study. Whether circRNAs are degraded in vivo by a similar mechanism still needs further

RNAi challenges this notion by demonstrating that RNA can also disrupt this flow of genetic information and prevent protein expression. Since 1998 the exact mechanism of RNAi has been thoroughly investigated, and it has been shown that the RNAi machinery, known as RNA-induced silencing complex (RISC), is activated when two fragments of RNA combine to form double-stranded RNA (Ramachandran et al., 2013). RISC then binds to one of these RNA fragments, which acts as a template allowing RISC to search for its complementary sequence on the target mRNA. As such, this fragment is known as the guide or ‘antisense’ strand (Wang et al., 2011). When RISC detects its matching sequence, the target mRNA is cleaved by certain nucleases or degraded by the cellular machinery, which effectively prevents translation of the transcript and silences expression of the protein (Wang et al., 2011). As described, the initial step in this process relies on the formation of an RNA duplex, which can occur in two pathways. Firstly, dsRNA can form in an endogenous manner due to the presence of non-coding regulatory RNA called micro (mi)RNA. miRNAs were discovered in the early 2000s as a result of Fire and Mello’s work, and they function as important post-transcriptional

Long non-coding RNAs (lncRNAs) make up a large portion of cellular transcripts in the mammalian transcriptome (Clark and Blackshaw 2014) . In all human transcripts, at least 10,000 non-coding RNAs are thought to be greater than 200 base pairs (bps) in length, with little to no coding capacity, and are therefore classed as lncRNAs (Zhu et al. 2013; Clark and Blackshaw 2014). In some cases, transcripts can reach up to 9kb but, on average, are around 500-1000 bases long (Chakraborty et al. 2014). For an RNA to be classed as long non-coding RNA, open reading frames (ORFs) must be no longer than 100 amino acids, although some studies have

Different approaches have been developed in order to understand the function of these genes. The first methods involved random integration of short hairpin RNAs (shRNAs) or short inhibitory RNAs (siRNAs) to inhibit the production of a protein or overexpress it to gain insights about its function. 1

We have isolated EVs from the cin1 mutant, CIN1-S isoform mutant, and the WT strains and extracted exRNA. Since the cin1 mutant is drastically reduced in growth, 3x more culture volume than WT was required to obtain an exRNA yield comparable to that of others. exRNA was submitted to the Beijing Genome Institute (BGI, Shenzhen, China) for cDNA synthesis and RNA-Seq. Approximately 100 ng cDNA from each strain was subjected to Illumina deep sequencing using BGISEQ-500 technology. A close to 27 million clean sRNA reads each was obtained from all three strains. The length of sRNA is between 17 nt and 33 nt with most being 21, 22, and 23 nt. The initial sRNA annotation provided by BGI suggested that approximately 96% sRNA sequences were mapped

On the evolutionary scale biological complexity is heavily increased, without any significant change in the number of protein coding genes whereas non-coding portion of the genome has substantially undergone change. It is now well accepted fact that on increasing evolutionary levels, majority of the genome gives rise to non-coding RNAs (ncRNAs) that led to the increased biological complexity. By halting the central dogma – the flow of information from DNA to RNA to protein, ncRNAs have gained central attention. Recent advancement in the next generation sequencing (NGS) technology allowed discovery of several classes of ncRNAs. lncRNAs are the largest class of ncRNAs, by convention longer than 200 nucleotides in length.

In recent years, the next-generation sequencing technology have been used as an powerfull tool by researchers to study cancer data, including cancer diagnosis, therapy, and prognosis. This technology enabled researchers to investigate all human genome transcripts, including non-coding versions. Recent investigations have revealed the association of non-coding RNAs (ncRNAs), including miRNAs and lncRNAs, with cancer pathogenesis, prognosis, survival rate and uses of these ncRNAs as cancer biomarker[22-25].

As it has been suggested functional miRNA are present above 0.1% of the total reads. A 0.1 % cut-off in was applied in all cell lines, for selection of functionally relevant miRNAs in our analysis [91, 101]. In comparison to U2932 and U2932-EBV, our data show that 52 out of 68 miRNA in total cellular miRNA profile, and 49 out of 57 miRNA in Ago2 miRNAs profile, even though at different ratios but were present in both U2932 and U2932-EBV cells(appendix x). In comparison of SUDHL5 and SUDHL5-EBV cells, we found that 55 out of 84 miRNA, in total cellular miRNA profile, and 46 out of 73 miRNA, in Ago2 miRNAs profile, even though at different ratios but were present in both SUDHL5 and SUDHL5-EBV cells .

RNAs were extracted from OLs or cerebral cortex of P2 ICR mice using the miRNAeasy Mini Kit (QIAGEN Inc. Valencia, CA). Concentration of RNAs from each sample was determined using NanoDrop ND-1000 spectrophotometer (Thermo Scientific). Equal amount of RNAs were then used as template to transcribe cDNA using a High capacity cDNA reverse transcription kit (Life Technologies).