Post-transcriptional regulation of human cathepsin L expression

2011 ◽  
Vol 392 (5) ◽  
Author(s):  
Shivani Mittal ◽  
Riyaz A. Mir ◽  
Shyam S. Chauhan
Keyword(s):  
Splice Variants ◽  
Cathepsin L ◽  
Live Cells ◽  
Luciferase Reporter ◽  
Entry Site ◽  
Lower Efficiency ◽  
Internal Ribosome Entry ◽  
Lysosomal Protease ◽  
Human Cathepsin

AbstractThe expression of cathepsin L, a lysosomal protease, is known to be elevated in cancer and other pathologies. Multiple splice variants of human cathepsin L with variable 5′UTRs exist, which encode for the same protein. Previously we have observed that variant hCATL A (bearing the longest 5′UTR) was translatedin vitrowith significantly lower efficiency than variant hCATL AIII (bearing the shortest 5′UTR). Contrary to these findings, results of the present study reveal that in cancer cells, hCATL A mRNA exhibits higher translatability in spite of having lower stability than AIII. This is the first report demonstrating a highly contrasting trend in translation efficiencies of hCATL variants in rabbit reticulocytes and live cells. Expression from chimeric mRNAs containing 5′UTRs of A or AIII upstream to luciferase reporter cDNA established the A UTR to be the sole determinant for this effect. Transient transfections of bicistronic plasmids and mRNAs confirmed the presence of a functional Internal Ribosome Entry Site in this UTR. Our data suggest that differential stability and translation initiation modes mediated by the 5′UTRs of human cathepsin L variants are involved in regulating its expression.

Splice Variants of Human Cathepsin L mRNA Show Different Expression Rates

2001 ◽  
Vol 382 (11) ◽  
pp. 1583-1591 ◽  
Author(s):  
Abulizi Abudula ◽  
Winfried Rommerskirch ◽  
Ekkehard Weber ◽  
Dagmar Günther ◽  
Bernd Wiederanders
Keyword(s):  
Malignant Tumors ◽  
Splice Variants ◽  
Cathepsin L ◽  
Mrna Levels ◽  
Kidney Tumors ◽  
Exon 1 ◽  
Expression Rate ◽  
Human Cathepsin

Abstract Human cathepsin L (hCATL) mRNA occurs in vivo in at least three splice variants. They differ in the length of exon 1, which comprises 278 nucleotides (hCATLA), 188 nucleotides (hCATLA2) and 132 nucleotides (hCATLA3), respectively. We describe here the shortest variant for the first time. This form is predominant in all tissues and cells examined so far, including malignant tumors. We studied the expression rate of the three mRNA variants in order to explain why malignant kidney tumors show low cathepsin L activity despite of high mRNA levels. The variant hCATLA3 showed the highest expression rate in vitro and in vivo. Based on these results, we suggest a cisacting element on human cathepsin L mRNA which can be bound by a negative transacting regulator, thus leading to reduced expression rates.

scholarly journals Production and Application of Multicistronic Constructs for Various Human Disease Therapies

Pharmaceutics ◽  
2019 ◽  
Vol 11 (11) ◽  
pp. 580 ◽  
Author(s):  
Alisa A. Shaimardanova ◽  
Kristina V. Kitaeva ◽  
Ilmira I. Abdrakhmanova ◽  
Vladislav M. Chernov ◽  
Catrin S. Rutland ◽  
...  
Keyword(s):  
Human Diseases ◽  
Cellular Biology ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
New Methods ◽  
In Vivo Experiments ◽  
Mutant Genes ◽  
Methods Of Treatment

The development of multicistronic vectors has opened up new opportunities to address the fundamental issues of molecular and cellular biology related to the need for the simultaneous delivery and joint expression of several genes. To date, the examples of the successful use of multicistronic vectors have been described for the development of new methods of treatment of various human diseases, including cardiovascular, oncological, metabolic, autoimmune, and neurodegenerative disorders. The safety and effectiveness of the joint delivery of therapeutic genes in multicistronic vectors based on the internal ribosome entry site (IRES) and self-cleaving 2A peptides have been shown in both in vitro and in vivo experiments as well as in clinical trials. Co-expression of several genes in one vector has also been used to create animal models of various inherited diseases which are caused by mutations in several genes. Multicistronic vectors provide expression of all mutant genes, which allows the most complete mimicking disease pathogenesis. This review comprehensively discusses multicistronic vectors based on IRES nucleotide sequence and self-cleaving 2A peptides, including its features and possible application for the treatment and modeling of various human diseases.

scholarly journals La Autoantigen Is Necessary for Optimal Function of the Poliovirus and Hepatitis C Virus Internal Ribosome Entry Site In Vivo and In Vitro

2004 ◽  
Vol 24 (15) ◽  
pp. 6861-6870 ◽  
Author(s):  
Mauro Costa-Mattioli ◽  
Yuri Svitkin ◽  
Nahum Sonenberg
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Internal Ribosome Entry Site ◽  
Ribosomal Subunit ◽  
Dominant Negative ◽  
Dominant Negative Mutant ◽  
Entry Site ◽  
Internal Ribosome Entry

ABSTRACT Translation of poliovirus and hepatitis C virus (HCV) RNAs is initiated by recruitment of 40S ribosomes to an internal ribosome entry site (IRES) in the mRNA 5′ untranslated region. Translation initiation of these RNAs is stimulated by noncanonical initiation factors called IRES trans-activating factors (ITAFs). The La autoantigen is such an ITAF, but functional evidence for the role of La in poliovirus and HCV translation in vivo is lacking. Here, by two methods using small interfering RNA and a dominant-negative mutant of La, we demonstrate that depletion of La causes a dramatic reduction in poliovirus IRES function in vivo. We also show that 40S ribosomal subunit binding to HCV and poliovirus IRESs in vitro is inhibited by a dominant-negative form of La. These results provide strong evidence for a function of the La autoantigen in IRES-dependent translation and define the step of translation which is stimulated by La.

The Role of AKT and ERK in Regulating D-Cyclin Translation Inhibition and G1 Arrest in Multiple Myeloma Cells Treated with mTOR Inhibitors: Rationale for Combination Thereapy.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4795-4795
Author(s):  
Patrick J. Frost ◽  
YiJiang Shi ◽  
Carolyne Bardalaban ◽  
Bao Hoang ◽  
Alan Lichtenstein
Keyword(s):  
Multiple Myeloma ◽  
Mtor Inhibitors ◽  
G1 Arrest ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Myeloma Cells ◽  
Transfected Cells ◽  
P38 Inhibitor

Abstract In a previous study, we showed that heightened AKT activity sensitized multiple myeloma (MM) cells to the in vivo anti-tumor effects of CCI-779. To test the mechanism of AKT’s regulatory role, we studied isogenic U266 MM cell lines transfected with an activated AKT allele or empty vector. The AKT-transfected cells were markedly more sensitive to cytostasis induced in vitro by rapamycin or in vivo by CCI-779. In contrast, cells with quiescent AKT were completely resistant. The ability of rapamycin and CCI-779 to inhibit D-cyclin expression was also significantly greater in AKT-transfected MM cells and this was, in part, due to a greater ability to curtail cap-independent translation and internal ribosome entry site (IRES) activity of D-cyclin transcripts. As ERK/p38 activity can facilitate IRES-mediated translation of some transcripts, we investigated ERK/p38 as regulators of rapamycin sensitivity. AKT-transfected cells demonstrated significantly decreased ERK and p38 activity, suggesting their involvement. However, only an ERK inhibitor prevented D-cyclin IRES activity in resistant “low AKT” myeloma cells while a p38 inhibitor had no effect. Furthermore, the combination of rapamycin and the ERK inhibitor successfully sensitized myeloma cells to rapamycin in terms of down regulated D-cyclin protein expression and G1 arrest. These data support a scenario where ERK facilitates D-cyclin IRES function and heightened AKT activity down regulates this ERK-dependent phenomenon. Thus ERK and AKT activity are potential predictors of responsiveness to mTOR inhibitors.

scholarly journals Virological characterization of the hepatitis C virus JFH-1 strain in lymphocytic cell lines

2008 ◽  
Vol 89 (7) ◽  
pp. 1587-1592 ◽  
Author(s):  
Kyoko Murakami ◽  
Toshiro Kimura ◽  
Motonao Osaki ◽  
Koji Ishii ◽  
Tatsuo Miyamura ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Cell Lines ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Culture Systems ◽  
Polyprotein Processing ◽  
Lymphocytic Cell

While hepatocytes are the major site of hepatitis C virus (HCV) infection, a number of studies have suggested that HCV can replicate in lymphocytes. However, in vitro culture systems to investigate replication of HCV in lymphocytic cells are severely limited. Robust HCV culture systems have been established using the HCV JFH-1 strain and Huh-7 cells. To gain more insights into the tissue tropism of HCV, we investigated the infection, replication, internal ribosome entry site (IRES)-dependent translation and polyprotein processing of the HCV JFH-1 strain in nine lymphocytic cell lines. HCV JFH-1 failed to infect lymphocytes and replicate, but exhibited efficient polyprotein processing and IRES-dependent translation in lymphocytes as well as in Huh-7 cells. Our results suggest that lymphocytic cells can support HCV JFH-1 translation and polyprotein processing, but may lack some host factors essential for HCV JFH-1 infection and replication.

scholarly journals Effect of NS3 and NS5B proteins on classical swine fever virus internal ribosome entry site-mediated translation and its host cellular translation

2008 ◽  
Vol 89 (4) ◽  
pp. 994-999 ◽  
Author(s):  
Ming Xiao ◽  
Yan Bai ◽  
Hui Xu ◽  
Xiaolu Geng ◽  
Jun Chen ◽  
...  
Keyword(s):  
Internal Ribosome Entry Site ◽  
Classical Swine Fever ◽  
Rna Helicase ◽  
Helicase Domain ◽  
Dependent Manner ◽  
Helicase Activity ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Cellular Translation

A full-length NS3 (NS3F) and a truncated NS3 protein (NS3H) with an RNA helicase domain possess RNA helicase activity. Using an in vitro system with a monocistronic reporter RNA or DNA, containing the CSFV 5′-UTR, we observed that both NS3F and NS3H enhanced internal ribosome entry site (IRES)-mediated and cellular translation in a dose-dependent manner, but NS3 protease (NS3P) that lacks a helicase domain did not. NS3F was stronger than NS3H in promoting both translations. These results showed that viral RNA helicase could promote viral and cellular translation, and higher RNA helicase activity might be more efficient. The NS5B protein, the viral replicase, did not significantly affect the IRES-directed or cellular translation alone. NS5B significantly enhanced the stimulative effect of NS3F on both IRES-mediated and cellular translation, but did not affect that of NS3H or NS3P. This suggests that NS5B and NS3 interact via the protease domain during the enhancement of translation.

scholarly journals Mislocalization of CDK11/PITSLRE, a regulator of the G2/M phase of the cell cycle, in Alzheimer disease

2011 ◽  
Vol 16 (3) ◽  
Author(s):  
Vladan Bajić ◽  
Bo Su ◽  
Hyoung-Gon Lee ◽  
Wataru Kudo ◽  
Sandra Siedlak ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Alzheimer Disease ◽  
Expression Patterns ◽  
Vital Role ◽  
Dependent Manner ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Cellular Processes ◽  
M Phase

AbstractPost-mitotic neurons are typically terminally differentiated and in a quiescent status. However, in Alzheimer disease (AD), many neurons display ectopic re-expression of cell cycle-related proteins. Cyclin-dependent kinase 11 (CDK11) mRNA produces a 110-kDa protein (CDK11p110) throughout the cell cycle, a 58-kDa protein (CDK11p58) that is specifically translated from an internal ribosome entry site and expressed only in the G2/M phase of the cell cycle, and a 46-kDa protein (CDK11p46) that is considered to be apoptosis specific. CDK11 is required for sister chromatid cohesion and the completion of mitosis. In this study, we found that the expression patterns of CDK11 vary such that cytoplasmic CDK11 is increased in AD cellular processes, compared to a pronounced nuclear expression pattern in most controls. We also investigated the effect of amyloid precursor protein (APP) on CDK11 expression in vitro by using M17 cells overexpressing wild-type APP and APP Swedish mutant phenotype and found increased CDK11 expression compared to empty vector. In addition, amyloid-β25–35 resulted in increased CDK11 in M17 cells. These data suggest that CDK11 may play a vital role in cell cycle re-entry in AD neurons in an APP-dependent manner, thus presenting an intriguing novel function of the APP signaling pathway in AD.

scholarly journals Cancer-associated mRNAs regulated by the Helix-Loop-Helix motif of human EIF3A

2018 ◽  
Author(s):  
Marina Volegova ◽  
Jamie H.D. Cate
Keyword(s):  
Translation Initiation ◽  
Rna Binding ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Binding Motif ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Helix Loop Helix ◽  
Small Set

AbstractImproper regulation of translation initiation, a vital check-point of protein synthesis in the cell, has been linked to a number of cancers. Overexpression of protein subunits of eukaryotic translation initiation factor 3 (eIF3) has been associated with increased translation of mRNAs involved in cell proliferation. In addition to playing a major role in general translation initiation by serving as a scaffold for the assembly of translation initiation complexes, eIF3 regulates translation of specific cellular mRNAs and viral RNAs. Mutations in the N-terminal Helix-Loop-Helix (HLH) RNA-binding motif of the EIF3A subunit in eIF3 interfere with Hepatitis C Virus Internal Ribosome Entry Site (IRES) mediated translation initiationin vitro. Here we show that the EIF3A HLH motif controls translation of a small set of cellular transcripts enriched in oncogenic mRNAs, includingMYC. We also demonstrate that the HLH motif of EIF3A acts specifically on the 5’-UTR ofMYCmRNA and modulates the function of EIF4A1 on select transcripts during translation initiation. In Ramos lymphoma cell lines, which are dependent on MYC overexpression, mutations in the HLH motif greatly reduce MYC expression, impede proliferation and sensitize cells to anti-cancer compounds. These results reveal the potential of the EIF3A HLH motif in eIF3 as a promising chemotherapeutic target.SummaryThe Helix Loop Helix motif of EIF3A controls translation of a small set of oncogenic cellular transcripts, includingMYC, and modulates the function of translation initiation factor EIF4A1 during translation initiation on select mRNAs.

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