Giardiavirus internal ribosome entry site has an apparently unique mechanism of initiating translation.

Giardiavirus (GLV) utilizes an internal ribosome entry site (IRES) for translation initiation in the early branching eukaryote Giardia lamblia. Unlike most of the viral IRESs among higher eukaryotes, which localize primarily within the 5'-untranslated region (UTR), the GLV IRES comprises 253 nts of 5'UTR and the initial 264 nts in the open-reading-frame (ORF). To test if GLV IRES also functions in higher eukaryotic systems, we examined it in rabbit reticulocyte lysate (RRL) and found that it functions much less efficiently than the IRES from the Encephalomyocarditis virus (EMCV) or Cricket paralysis virus (CrPV). In contrast, both EMCV-IRES and CrPV-IRESs were inactive in transfected Giardia cells. Structure-function analysis indicated that only the stem-loop U5 from the 5'UTR and the stem-loop I plus the downstream box (Dbox) from the ORF of GLV IRES are required for limited IRES function in RRL. Edeine, a translation initiation inhibitor, did not significantly affect the function of GLV IRES in either RRL or Giardia, indicating that a pre-initiation complex is not required for GLV IRES-mediated translation initiation. However, the small ribosomal subunit purified from Giardia did not bind to GLV IRES, indicating that additional protein factors may be necessary. A member of the helicase family IBP1 and two known viral IRES binding proteins La autoantigen and SRp20 have been identified in Giardia that bind to GLV IRES in vitro. These three proteins could be involved in facilitating small ribosome recruitment for initiating translation

A La Autoantigen Homologue Is Required for the Internal Ribosome Entry Site Mediated Translation of Giardiavirus

Translation of Giardiavirus (GLV) mRNA is initiated at an internal ribosome entry site (IRES) in the viral transcript. The IRES localizes to a downstream portion of 5′ untranslated region (UTR) and a part of the early downstream coding region of the transcript. Recent studies indicated that the IRES does not require a pre-initiation complex to initiate translation but may directly recruit the small ribosome subunit with the help of a number of trans-activating protein factors. A La autoantigen homologue in the viral host Giardia lamblia, GlLa, was proposed as one of the potential trans-activating factors based on its specific binding to GLV-IRES in vitro. In this study, we further elucidated the functional role of GlLa in GLV-IRES mediated translation in Giardia by knocking down GlLa with antisense morpholino oligo, which resulted in a reduction of GLV-IRES activity by 40%. An over-expression of GlLa in Giardia moderately stimulated GLV-IRES activity by 20%. A yeast inhibitory RNA (IRNA), known to bind mammalian and yeast La autoantigen and inhibit Poliovirus and Hepatitis C virus IRES activities in vitro and in vivo, was also found to bind to GlLa protein in vitro and inhibited GLV-IRES function in vivo. The C-terminal domain of La autoantigen interferes with the dimerization of La and inhibits its function. An over-expression of the C-terminal domain (200–348aa) of GlLa in Giardia showed a dominant-negative effect on GLV-IRES activity, suggesting a potential inhibition of GlLa dimerization. HA tagged GlLa protein was detected mainly in the cytoplasm of Giardia, thus supporting a primary role of GlLa in translation initiation in Giardiavirus

Structural Elements in the 5′-Untranslated Region of Giardiavirus Transcript Essential for Internal Ribosome Entry Site-Mediated Translation Initiation

Translation of uncapped giardiavirus (GLV) mRNA in Giardia lamblia requires the presence of a 5′-untranslated region (5′-UTR) and a viral capsid coding region. We used dicistronic viral constructs to show that the downstream 253 nucleotides (nt) of the 5′-UTR plus the initial 264-nt capsid coding region constitute an internal ribosome entry site (IRES). Predicted secondary structures in the 253-nt 5′-UTR include stem-loops U3, U4a, U4b, U4c, and U5. Chemical and enzymatic probing analysis confirmed the presence of all predicted stem-loops except U4a. Disruption of stem-loop structures U3 and U5 by site-directed mutagenesis resulted in a drastic reduction in translation of a monocistronic viral transcript, which could be restored by compensatory sequence changes. Mutations disrupting stem-loops U4b and U4c do not exert an appreciable effect on translation, but certain sequences in the U4a region and in U4b do appear to play important roles in the IRES. Structural analysis also suggests that an 8-nt U3 loop sequence (nt 147 to 154) pairs with an 8-nt downstream sequence (nt 168 to 175) to form a pseudoknot. Disruption of this pseudoknot by mutagenesis resulted in a drastic reduction in translation, which could be restored by compensatory sequence changes. This study has defined the secondary structure in the 5′-UTR of the IRES. Together with the previous results, we have now completed analysis of the entire structure of GLV IRES and fully defined the functionally essential structural elements in it

Giardiavirus internal ribosome entry site has an apparently unique mechanism of initiating translation.

Giardiavirus (GLV) utilizes an internal ribosome entry site (IRES) for translation initiation in the early branching eukaryote Giardia lamblia. Unlike most of the viral IRESs among higher eukaryotes, which localize primarily within the 5'-untranslated region (UTR), the GLV IRES comprises 253 nts of 5'UTR and the initial 264 nts in the open-reading-frame (ORF). To test if GLV IRES also functions in higher eukaryotic systems, we examined it in rabbit reticulocyte lysate (RRL) and found that it functions much less efficiently than the IRES from the Encephalomyocarditis virus (EMCV) or Cricket paralysis virus (CrPV). In contrast, both EMCV-IRES and CrPV-IRESs were inactive in transfected Giardia cells. Structure-function analysis indicated that only the stem-loop U5 from the 5'UTR and the stem-loop I plus the downstream box (Dbox) from the ORF of GLV IRES are required for limited IRES function in RRL. Edeine, a translation initiation inhibitor, did not significantly affect the function of GLV IRES in either RRL or Giardia, indicating that a pre-initiation complex is not required for GLV IRES-mediated translation initiation. However, the small ribosomal subunit purified from Giardia did not bind to GLV IRES, indicating that additional protein factors may be necessary. A member of the helicase family IBP1 and two known viral IRES binding proteins La autoantigen and SRp20 have been identified in Giardia that bind to GLV IRES in vitro. These three proteins could be involved in facilitating small ribosome recruitment for initiating translation

Future wind speed trends in the Indian offshore region

Climate models help assess the future availability of wind speeds to extract wind power; however, these climate models are mathematical models that include uncertainty in wind forecasts. Finding an appropriate climate model for analyzing future wind power generation is critical. Therefore, in the present work, six Coordinated Regional Climate Downscaling Experiment-South Asia (CORDEX-SA) climate models and their ensembles were statistically examined in the Indian offshore region using in-situ buoy readings and ERA5 reanalysis data. Statistical parameters like correlation coefficient, bias, RMSD, and standard deviation are computed to examine the applicable model at buoy locations. With ERA5 wind speeds, the overlapping percentage of climate models is later analyzed for the Indian offshore regions. The ensemble model statistically outperforms individual climate models at buoy sites, with 77% overlap with ERA5 wind speeds in the offshore region. Further, the trends and cumulative variations in wind speeds are calculated for ensemble models under two emission scenarios RCP (Representative Concentration Pathway) 4.5 and RCP 8.5. In the future, the North-East (NE) zone will have the most advantageous change in wind speeds (0.21 to 8.68%), whereas the North-West (NW) area will have a negative cumulative change in wind speeds