Activation of the tumor suppressor p53 upon impairment of ribosome biogenesis

Errors in ribosome biogenesis can result in quantitative or qualitative defects in protein synthesis and conse- quently lead to improper execution of the genetic program and the development of speci fi c diseases. Evidence has accumulated over the last decade suggesting that perturbation of ribosome biogenesis triggers a p53- activating checkpointsignaling pathway,often referred to as the ribosome biogenesis stress checkpoint pathway. Although it was originally suggested that p53 has a pro minent role in preventing diseases by monitoring the fi delity of ribosome biogenesis, recent work has demonstrated that p53 activation upon impairment of ribosome biogenesis also mediates pathological manifestations in humans . Perturbations of ribosome biogenesis can trigger a p53-dependent checkpoint signaling pathway independent of DNA damage and the tumor suppressor ARF through inhibitory interactions of speci fi c ribosomal components with the p53 negative regulator, Mdm2. Here we review the recent advances made toward understanding of this newly-recognized checkpoint signaling pathway, its role in health and disease, and discuss possible future directions in this exciting research fi eld. This article is part of a Special Issue entitled: Role of the Nucleolus in Human Disease

Mutual protection of ribosomal proteins L5 and L11 from degradation is essential for p53 activation upon ribosomal biogenesis stress

Impairment of ribosomal biogenesis can activate the p53 protein independently of DNA damage. The ability of ribosomal proteins L5, L11, L23, L26, or S7 to bind Mdm2 and inhibit its ubiquitin ligase activity has been suggested as a critical step in p53 activation under these conditions. Here, we report that L5 and L11 are particularly important for this response. Whereas several other newly synthesized ribosomal proteins are degraded by proteasomes upon inhibition of Pol I activity by actinomycin D, L5 and L11 accumulate in the ribosome-free fraction where they bind to Mdm2. This selective accumulation of free L5 and L11 is due to their mutual protection from proteasomal degradation. Furthermore, the endogenous, newly synthesized L5 and L11 continue to be imported into nucleoli even after nucleolar disruption and colocalize with Mdm2, p53, and promyelocytic leukemia protein. This suggests that the disrupted nucleoli may provide a platform for L5- and L11-dependent p53 activation, implying a role for the nucleolus in p53 activation by ribosomal biogenesis stress. These findings may have important implications with respect to understanding the pathogenesis of diseases caused by impaired ribosome biogenesis

Mechanisms by which the p53 tumor suppressor mediates pathological manifestations in Rpl24-heterozygous mice

Ciljevi istraživanja: Povišen izražaj gena p53 i Sesn2 u Rpl24-heterozigotnim miševima ukazuje da p53 regulira njihov fenotip putem regulacije mTOR-a i makroautofagije vjerojatno putem Sesn2. Ciljevi su: 1. Odrediti uzrokuje li p53 patološki fenotip Rpl24-heterozigotnih miševa putem Sesn2. 2. Odrediti aktivnost i važnost makroautofagije u preživljavanju Rpl24-heterozigotnih miševa. 3. Razjasniti molekularne mehanizme putem kojih p53 i Sesn2 reguliraju patološke fenotipove u Rpl24-heterozigotnim embrijima. Materijali i metode: U istraživanju je korišten miš s heterozigotnom mutacijom u genu za Rpl24, koja rezultira specifičnim fenotipom ovih miševa: slijepoćom, abnormalnosti skeleta, smanjenom masom i zavijenim repom (Bst, engl. Belly Spot and Tail). Izražaj Sesn2 utvrđen je u Rpl24-heterozigotnim embrijima na E10.5 korištenjem kvantitativne reakcije lančanom polimerazom u realnom vremenu (RT-PCR, engl. real time polymerase chain reaction). Da bih testirala mogućnost da p53 uzrokuje patološke fenotipove Rpl24-heterozigotnih miševa putem Sesn2, genetički sam u njima inaktivirala Sesn2. Da bih razlučila potencijalnu važnost makroautofagije i mTOR signalnog puta u Sesn2-ovisnom patološkom fenotipu Rpl24-heterozigotnih miševa, inaktivirala sam jedan alel Atg5, ključnog gena za makroautofagiju. Molekularne mehanizme regulacije preživljavanja Rpl24-heterozigotnih miševa razjasnila sam u modelu gladovanja u ranom poslijenatalnom periodu korištenjem Rpl24-heterozigotnih miševa u kojima je genetički inaktiviran p53, Sesn2 ili jedan alel Atg5. Rezultati: Dokazano je da p53 uzrokuje kongenitalne malformacije u Rpl24-heterozigotnim miševima putem svog ciljnog gena, Sesn2, te da p53 omogućuje preživljavanje Rpl24-heterozigotnih miševa u ranom poslijenatalnom periodu putem regulacije makroautofagije, koja najvjerojatnije razgradnjom staničnih komponenti osigurava supstrate za sintezu glukoze neophodne za njihovo preživljavanje. p53 i Sesn2 neophodni su za preživljavanje Rpl24-heterozigotnih miševa u ranom poslijenatalnom periodu gladovanja. Međutim, mehanizmi putem kojih p53 i Sesn2 potiču preživljavanje Rpl24-heterozigotnih miševa barem su djelomično različiti. Zaključci: Dokazano je da p53 uzrokuje kongenitalne malformacije u Rpl24-heterozigotnim miševima putem svog ciljnog gena, Sesn2, te su razjašnjeni molekularni mehanizmi kojima Sesn2 i p53 reguliraju patološke fenotipove u Rpl24-heterozigotnim miševima. Naši rezultati mogu pomoći razumijevanju mehanizama putem kojih p53 uzrokuje patološke promjene u ribosomopatijama, omogućiti bolji uvid u njihovu patogenezu i rezultirati otkrićem potencijalnih ciljeva za liječenje tih bolesti.Title: Mechanisms by which the p53 tumor suppressor mediates pathological manifestations in Rpl24-heterozygous mice Objectives: Upregulation of p53 and Sesn2 in Rpl24-heterozygous mice suggests that p53 regulates pathological manifestations of these mice through Sesn2-dependent regulation of autophagy and mTOR signalling pathway. Our objectives are: 1. To determine if p53-dependent induction of Sesn2 causes pathological phenotype of Rpl24-heterozygous mice 2. To determine if autophagy is activated and important for the survival of Rpl24-heterozygous mice 3. To determine p53- and Sesn2-dependent molecular mechanisms of regulation of pathological phenotypes of Rpl24-heterozygous mice. Material and Methods: Spontaneous heterozygous mutation in one allele of ribosomal protein l24 gene (Rpl24) in mice results in defects of the eye, skeleton, mass and coat pigmentation (Bst, Belly Spot and Tail). Sesn2 expression level was determined in Rpl24-heterozygous embryos at E10.5 using quantitative polymerase chain reaction (RT-PCR). To test the possibility that p53-dependent induction of Sesn2 plays a role in pathological phenotypes of Rpl24-heterozygous mice, I genetically inactivated Sesn2 in these mice. To dissect the importance of autophagy and mTOR signalling pathway in Sesn2-dependent pathological phenotype of Rpl24-heterozygous mice, I genetically inactivated one allele of Atg5, important autophagy gene. Molecular mechanisms of survival regulation in Rpl24-heterozygous mice were elucidated in the model of survival under fasting conditions using Rpl24-heterozygous mice with p53, Sesn2 or Atg5 inactivation. Results: We demonstrate the key role of p53 target gene, Sesn2, in pathological phenotype of Rpl24-heterozygous mice. We show that p53 promotes survival of these mice in early postnatal period through regulation of autophagy, which provides substrates for gluconeogenesis necessary for survival of Rpl24-heterozygous mice. Both, p53 and Sesn2, are indispensable for survival of Rpl24-heterozygous mice in the early postnatal starvation period, but the mechanisms through which these two genes regulate survival of these mice are at least partially different. Conclusions: We demonstrate the key role of p53 target gene, Sesn2, in pathological phenotype of Rpl24-heterozygous mice, and show the molecular mechanisms by which p53 and Sesn2 regulate these phenotypes. Our findings may have important implications for understanding p53-dependent mechanisms underlying the pathogenesis of ribosomopathies and could result in the discovery of potential targets for treatment of these diseases

Mechanisms by which the p53 tumor suppressor mediates pathological manifestations in Rpl24-heterozygous mice

Ciljevi istraživanja: Povišen izražaj gena p53 i Sesn2 u Rpl24-heterozigotnim miševima ukazuje da p53 regulira njihov fenotip putem regulacije mTOR-a i makroautofagije vjerojatno putem Sesn2. Ciljevi su: 1. Odrediti uzrokuje li p53 patološki fenotip Rpl24-heterozigotnih miševa putem Sesn2. 2. Odrediti aktivnost i važnost makroautofagije u preživljavanju Rpl24-heterozigotnih miševa. 3. Razjasniti molekularne mehanizme putem kojih p53 i Sesn2 reguliraju patološke fenotipove u Rpl24-heterozigotnim embrijima. Materijali i metode: U istraživanju je korišten miš s heterozigotnom mutacijom u genu za Rpl24, koja rezultira specifičnim fenotipom ovih miševa: slijepoćom, abnormalnosti skeleta, smanjenom masom i zavijenim repom (Bst, engl. Belly Spot and Tail). Izražaj Sesn2 utvrđen je u Rpl24-heterozigotnim embrijima na E10.5 korištenjem kvantitativne reakcije lančanom polimerazom u realnom vremenu (RT-PCR, engl. real time polymerase chain reaction). Da bih testirala mogućnost da p53 uzrokuje patološke fenotipove Rpl24-heterozigotnih miševa putem Sesn2, genetički sam u njima inaktivirala Sesn2. Da bih razlučila potencijalnu važnost makroautofagije i mTOR signalnog puta u Sesn2-ovisnom patološkom fenotipu Rpl24-heterozigotnih miševa, inaktivirala sam jedan alel Atg5, ključnog gena za makroautofagiju. Molekularne mehanizme regulacije preživljavanja Rpl24-heterozigotnih miševa razjasnila sam u modelu gladovanja u ranom poslijenatalnom periodu korištenjem Rpl24-heterozigotnih miševa u kojima je genetički inaktiviran p53, Sesn2 ili jedan alel Atg5. Rezultati: Dokazano je da p53 uzrokuje kongenitalne malformacije u Rpl24-heterozigotnim miševima putem svog ciljnog gena, Sesn2, te da p53 omogućuje preživljavanje Rpl24-heterozigotnih miševa u ranom poslijenatalnom periodu putem regulacije makroautofagije, koja najvjerojatnije razgradnjom staničnih komponenti osigurava supstrate za sintezu glukoze neophodne za njihovo preživljavanje. p53 i Sesn2 neophodni su za preživljavanje Rpl24-heterozigotnih miševa u ranom poslijenatalnom periodu gladovanja. Međutim, mehanizmi putem kojih p53 i Sesn2 potiču preživljavanje Rpl24-heterozigotnih miševa barem su djelomično različiti. Zaključci: Dokazano je da p53 uzrokuje kongenitalne malformacije u Rpl24-heterozigotnim miševima putem svog ciljnog gena, Sesn2, te su razjašnjeni molekularni mehanizmi kojima Sesn2 i p53 reguliraju patološke fenotipove u Rpl24-heterozigotnim miševima. Naši rezultati mogu pomoći razumijevanju mehanizama putem kojih p53 uzrokuje patološke promjene u ribosomopatijama, omogućiti bolji uvid u njihovu patogenezu i rezultirati otkrićem potencijalnih ciljeva za liječenje tih bolesti.Title: Mechanisms by which the p53 tumor suppressor mediates pathological manifestations in Rpl24-heterozygous mice Objectives: Upregulation of p53 and Sesn2 in Rpl24-heterozygous mice suggests that p53 regulates pathological manifestations of these mice through Sesn2-dependent regulation of autophagy and mTOR signalling pathway. Our objectives are: 1. To determine if p53-dependent induction of Sesn2 causes pathological phenotype of Rpl24-heterozygous mice 2. To determine if autophagy is activated and important for the survival of Rpl24-heterozygous mice 3. To determine p53- and Sesn2-dependent molecular mechanisms of regulation of pathological phenotypes of Rpl24-heterozygous mice. Material and Methods: Spontaneous heterozygous mutation in one allele of ribosomal protein l24 gene (Rpl24) in mice results in defects of the eye, skeleton, mass and coat pigmentation (Bst, Belly Spot and Tail). Sesn2 expression level was determined in Rpl24-heterozygous embryos at E10.5 using quantitative polymerase chain reaction (RT-PCR). To test the possibility that p53-dependent induction of Sesn2 plays a role in pathological phenotypes of Rpl24-heterozygous mice, I genetically inactivated Sesn2 in these mice. To dissect the importance of autophagy and mTOR signalling pathway in Sesn2-dependent pathological phenotype of Rpl24-heterozygous mice, I genetically inactivated one allele of Atg5, important autophagy gene. Molecular mechanisms of survival regulation in Rpl24-heterozygous mice were elucidated in the model of survival under fasting conditions using Rpl24-heterozygous mice with p53, Sesn2 or Atg5 inactivation. Results: We demonstrate the key role of p53 target gene, Sesn2, in pathological phenotype of Rpl24-heterozygous mice. We show that p53 promotes survival of these mice in early postnatal period through regulation of autophagy, which provides substrates for gluconeogenesis necessary for survival of Rpl24-heterozygous mice. Both, p53 and Sesn2, are indispensable for survival of Rpl24-heterozygous mice in the early postnatal starvation period, but the mechanisms through which these two genes regulate survival of these mice are at least partially different. Conclusions: We demonstrate the key role of p53 target gene, Sesn2, in pathological phenotype of Rpl24-heterozygous mice, and show the molecular mechanisms by which p53 and Sesn2 regulate these phenotypes. Our findings may have important implications for understanding p53-dependent mechanisms underlying the pathogenesis of ribosomopathies and could result in the discovery of potential targets for treatment of these diseases

Herpes Simplex Virus 1 Deregulation of Host MicroRNAs

Viruses utilize microRNAs (miRNAs) in a vast variety of possible interactions and mechanisms, apparently far beyond the classical understanding of gene repression in humans. Likewise, herpes simplex virus 1 (HSV-1) expresses numerous miRNAs and deregulates the expression of host miRNAs. Several HSV-1 miRNAs are abundantly expressed in latency, some of which are encoded antisense to transcripts of important productive infection genes, indicating their roles in repressing the productive cycle and/or in maintenance/reactivation from latency. In addition, HSV-1 also exploits host miRNAs to advance its replication or repress its genes to facilitate latency. Here, we discuss what is known about the functional interplay between HSV-1 and the host miRNA machinery, potential targets, and the molecular mechanisms leading to an efficient virus replication and spread

CD8 T Cell Vaccines and a Cytomegalovirus-Based Vector Approach

The twentieth century witnessed a huge expansion in the number of vaccines used with great success in combating diseases, especially the ones caused by viral and bacterial pathogens. Despite this, several major public health threats, such as HIV, tuberculosis, malaria, and cancer, still pose an enormous humanitarian and economic burden. As vaccines based on the induction of protective, neutralizing antibodies have not managed to effectively combat these diseases, in recent decades, the focus has increasingly shifted towards the cellular immune response. There is substantial evidence demonstrating CD8 T cells as key players in the protection not only against many viral and bacterial pathogens, but also in the fight against neoplastic cells. Here, we present arguments for CD8 T cells to be considered as promising candidates for vaccine targeting. We discuss the heterogeneity of CD8 T cell populations and their contribution in the protection of the host. We also outline several strategies of using a common human pathogen, cytomegalovirus, as a vaccine vector since accumulated data strongly suggest it represents a promising approach to the development of novel vaccines against both pathogens and tumors

Što je rak?

U sklopu ovogodišnje Noći istraživača, manifestacije financirane u okviru programa Horizon Europe koja ima za cilj približiti znanost i istraživanje javnosti, znanstvenici iz Centra za Proteomiku i Zavoda za Histologiju pripremili su 3 slikovnice naslova: „Što je rak?“, „Što su protutijela“ i „Što rade znanstvenici“. Navedene slikovnice imaju za cilj na jednostavan i razumljiv način objasniti djeci i mladima osnovne koncepte koji se tiču biologije raka, protutijela kao alata za istraživanje ili terapiju, te znanstvenika kao profesije koja sudjeluje u „borbi“ protiv raka. Naime, djeca, čak i ako nisu u stanju razumjeti rak, posebno su pažljiva i jako pogođena dijagnozom raka, a istraživanja pokazuju da otvorenost i iskrenost pomažu djeci da se nose s problemima. Osnovne informacije o raku pokrivene su u prvoj slikovnici “Što je rak?”, autorice dr.sc. Paole Kučan Brlić, koja je namijenjena djeci predškolske dobi i nižih razreda osnovne škole. Druga slikovnica, “Što su antitijela?”, autorice Dore Gašparini, dr.med., namijenjena je starijoj djeci te objašnjava kako funkcioniraju protutijela i kako se ona mogu koristiti za liječenje raka ili u znanstvenim istraživanjima. Zadnja slikovnica, „Što rade znanstvenici“, autorica dr.sc. Jelene Železnjak i dr.sc. Marije Mazor, predstavlja i promovira znanstvenike kao profesiju koja se, između ostalog, bavi istraživanjem i liječenjem raka, te objašnjava kako se postaje znanstvenik. Uz navedene autorice, u pripremi ovih slikovnica za tisak, sudjelovali su i ostali članovi tima Noći istraživača: izv. prof. dr. sc. Vanda Juranić Lisnić, dr. sc. Maja Cokarić Brdovčak, Cristina Paulović i Fran Krstanović. Sakrij dio sažetk

Što su protutijela?

U sklopu ovogodišnje Noći istraživača, manifestacije financirane u okviru programa Horizon Europe koja ima za cilj približiti znanost i istraživanje javnosti, znanstvenici iz Centra za Proteomiku i Zavoda za Histologiju pripremili su 3 slikovnice naslova: „Što je rak?“, „Što su protutijela“ i „Što rade znanstvenici“. Navedene slikovnice imaju za cilj na jednostavan i razumljiv način objasniti djeci i mladima osnovne koncepte koji se tiču biologije raka, protutijela kao alata za istraživanje ili terapiju, te znanstvenika kao profesije koja sudjeluje u „borbi“ protiv raka. Naime, djeca, čak i ako nisu u stanju razumjeti rak, posebno su pažljiva i jako pogođena dijagnozom raka, a istraživanja pokazuju da otvorenost i iskrenost pomažu djeci da se nose s problemima. Osnovne informacije o raku pokrivene su u prvoj slikovnici “Što je rak?”, autorice dr.sc. Paole Kučan Brlić, koja je namijenjena djeci predškolske dobi i nižih razreda osnovne škole. Druga slikovnica, “Što su antitijela?”, autorice Dore Gašparini, dr.med., namijenjena je starijoj djeci te objašnjava kako funkcioniraju protutijela i kako se ona mogu koristiti za liječenje raka ili u znanstvenim istraživanjima. Zadnja slikovnica, „Što rade znanstvenici“, autorica dr.sc. Jelene Železnjak i dr.sc. Marije Mazor, predstavlja i promovira znanstvenike kao profesiju koja se, između ostalog, bavi istraživanjem i liječenjem raka, te objašnjava kako se postaje znanstvenik. Uz navedene autorice, u pripremi ovih slikovnica za tisak, sudjelovali su i ostali članovi tima Noći istraživača: izv. prof. dr. sc. Vanda Juranić Lisnić, dr. sc. Maja Cokarić Brdovčak, Cristina Paulović i Fran Krstanović