scholarly journals The Absence of Mitochondrial Thioredoxin 2 Causes Massive Apoptosis, Exencephaly, and Early Embryonic Lethality in Homozygous Mice

2003 ◽  
Vol 23 (3) ◽  
pp. 916-922 ◽  
Author(s):  
Larisa Nonn ◽  
Ryan R. Williams ◽  
Robert P. Erickson ◽  
Garth Powis
Keyword(s):  
Nuclear Gene ◽  
Embryonic Lethality ◽  
Mouse Development ◽  
Redox Protein ◽  
Early Embryonic Lethality ◽  
Homozygous Mutant ◽  
Thioredoxin 2 ◽  
Massive Apoptosis

ABSTRACT Thioredoxin 2 (Trx-2) is a small redox protein containing the thioredoxin active site Trp-Cys-Gly-Pro-Cys that is localized to the mitochondria by a mitochondrial leader sequence and encoded by a nuclear gene (Trx-2). Trx-2 plays an important role in cell viability and the regulation of apoptosis in vitro. To investigate the role of Trx-2 in mouse development, we studied the phenotype of mice that have the Trx-2 gene silenced by mutational insertion. Homozygous mutant embryos do not survive to birth and die after implantation at Theiler stage 15/16. The homozygous mutant embryos display an open anterior neural tube and show massively increased apoptosis at 10.5 days postcoitus and are not present by 12.5 days postcoitus. The timing of the embryonic lethality coincides with the maturation of the mitochondria, since they begin oxidative phosphorylation during this stage of embryogenesis. In addition, embryonic fibroblasts cultured from homozygous Trx-2-null embryos were not viable. Heterozygous mice are fertile and have no discernible phenotype visible by external observation, despite having decreased Trx-2 mRNA and protein. These results show that the mitochondrial redox protein Trx-2 is required for normal development of the mouse embryo and for actively respiring cells.

Defects in mesoderm, neural tube and vascular development in mouse embryos lacking fibronectin

Development ◽  
1993 ◽  
Vol 119 (4) ◽  
pp. 1079-1091 ◽  
Author(s):  
E.L. George ◽  
E.N. Georges-Labouesse ◽  
R.S. Patel-King ◽  
H. Rayburn ◽  
R.O. Hynes
Keyword(s):  
Mutant Allele ◽  
Mouse Embryos ◽  
Neural Tubes ◽  
Early Embryonic Lethality ◽  
Homozygous Mutant ◽  
Anterior Posterior ◽  
Embryonic Vessels

To examine the role of fibronectin in vivo, we have generated mice in which the fibronectin gene is inactivated. Heterozygotes have one half normal levels of plasma fibronectin, yet appear normal. When homozygous, the mutant allele causes early embryonic lethality, proving that fibronectin is required for embryogenesis. However, homozygous mutant embryos implant and initiate gastrulation normally including extensive mesodermal movement. Neural folds also form but the mutant embryos subsequently display shortened anterior-posterior axes, deformed neural tubes and severe defects in mesodermally derived tissues. Notochord and somites are absent; the heart and embryonic vessels are variable and deformed, and the yolk sac, extraembryonic vasculature and amnion are also defective. These abnormalities can be interpreted as arising from fundamental deficits in mesodermal migration, adhesion, proliferation or differentiation as a result of the absence of fibronectin. The nature of these embryonic defects leads to reevaluation of suggested roles for fibronectin during early development based on results obtained in vitro and in embryos of other species.

scholarly journals Rpn10-Mediated Degradation of Ubiquitinated Proteins Is Essential for Mouse Development

2007 ◽  
Vol 27 (19) ◽  
pp. 6629-6638 ◽  
Author(s):  
Jun Hamazaki ◽  
Katsuhiro Sasaki ◽  
Hiroyuki Kawahara ◽  
Shin-ichi Hisanaga ◽  
Keiji Tanaka ◽  
...  
Keyword(s):  
Splice Variants ◽  
Biological Significance ◽  
Embryonic Lethality ◽  
26S Proteasome ◽  
Mouse Development ◽  
Developmentally Regulated ◽  
Ubiquitinated Proteins ◽  
Early Embryonic Lethality ◽  
Von Willebrand

ABSTRACT Rpn10 is a subunit of the 26S proteasome that recognizes polyubiquitinated proteins. The importance of Rpn10 in ubiquitin-mediated proteolysis is debatable, since a deficiency of Rpn10 causes different phenotypes in different organisms. To date, the role of mammalian Rpn10 has not been examined genetically. Moreover, vertebrates have five splice variants of Rpn10 whose expressions are developmentally regulated, but their biological significance is not understood. To address these issues, we generated three kinds of Rpn10 mutant mice. Rpn10 knockout resulted in early-embryonic lethality, demonstrating the essential role of Rpn10 in mouse development. Rpn10a knock-in mice, which exclusively expressed the constitutive type of Rpn10 and did not express vertebrate-specific variants, grew normally, indicating that Rpn10 diversity is not essential for conventional development. Mice expressing the N-terminal portion of Rpn10, which contained a von Willebrand factor A (VWA) domain but lacked ubiquitin-interacting motifs (Rpn10ΔUIM), also exhibited embryonic lethality, suggesting the important contribution of UIM domains to viability, but survived longer than Rpn10-null mice, consistent with a “facilitator” function of the VWA domain. Biochemical analysis of the Rpn10ΔUIM liver showed specific impairment of degradation of ubiquitinated proteins. Our results demonstrate that Rpn10-mediated degradation of ubiquitinated proteins, catalyzed by UIMs, is indispensable for mammalian life.

Transposon-Disruption of a Maize Nuclear Gene, tha1, Encoding a Chloroplast SecA Homologue: In Vivo Role of cp-SecA in Thylakoid Protein Targeting

Genetics ◽  
1997 ◽  
Vol 145 (2) ◽  
pp. 467-478 ◽  
Author(s):  
Rodger Voelker ◽  
Janet Mendel-Hartvig ◽  
Alice Barkan
Keyword(s):  
Protein Targeting ◽  
Sequence Similarity ◽  
Nuclear Gene ◽  
Cytochrome F ◽  
Transposon Insertion ◽  
Nuclear Mutant ◽  
Strong Sequence Similarity

A nuclear mutant of maize, tha1, which exhibited defects in the translocation of proteins across the thylakoid membrane, was described previously. A transposon insertion at the tha1 locus facilitated the cloning of portions of the tha1 gene. Strong sequence similarity with secA genes from bacteria, pea and spinach indicates that tha1 encodes a SecA homologue (cp-SecA). The tha1-ref allele is either null or nearly so, in that tha1 mRNA is undetectable in mutant leaves and cp-SecA accumulation is reduced ≥40-fold. These results, in conjunction with the mutant phenotype described previously, demonstrate that cp-SecA functions in vivo to facilitate the translocation of OEC33, PSI-F and plastocyanin but does not function in the translocation of OEC23 and OEC16. Our results confirm predictions for cp-Sed function made from the results of in vitro experiments and establish several new functions for cp-SecA, including roles in the targeting of a chloroplast-encoded protein, cytochrome f, and in protein targeting in the etioplast, a nonphotosynthetic plastid type. Our finding that the accumulation of properly targeted plastocyanin and cytochrome f in tha1-ref thylakoid membranes is reduced only a few-fold despite the near or complete absence of cp-SecA suggests that cp-SecA facilitates but is not essential in vivo for their translocation across the membrane.

scholarly journals Redundant Roles of Tead1 and Tead2 in Notochord Development and the Regulation of Cell Proliferation and Survival

2008 ◽  
Vol 28 (10) ◽  
pp. 3177-3189 ◽  
Author(s):  
Atsushi Sawada ◽  
Hiroshi Kiyonari ◽  
Kanako Ukita ◽  
Noriyuki Nishioka ◽  
Yu Imuta ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Paraxial Mesoderm ◽  
Mouse Development ◽  
Lateral Plate ◽  
Growth Defects ◽  
Mouse Mutants ◽  
Severe Growth

ABSTRACT Four members of the TEAD/TEF family of transcription factors are expressed widely in mouse embryos and adult tissues. Although in vitro studies have suggested various roles for TEAD proteins, their in vivo functions remain poorly understood. Here we examined the role of Tead genes by generating mouse mutants for Tead1 and Tead2. Tead2 −/− mice appeared normal, but Tead1 −/−; Tead2 −/− embryos died at embryonic day 9.5 (E9.5) with severe growth defects and morphological abnormalities. At E8.5, Tead1 −/−; Tead2 −/− embryos were already small and lacked characteristic structures such as a closed neural tube, a notochord, and somites. Despite these overt abnormalities, differentiation and patterning of the neural plate and endoderm were relatively normal. In contrast, the paraxial mesoderm and lateral plate mesoderm were displaced laterally, and a differentiated notochord was not maintained. These abnormalities and defects in yolk sac vasculature organization resemble those of mutants for Yap, which encodes a coactivator of TEAD proteins. Moreover, we demonstrated genetic interactions between Tead1 and Tead2 and Yap. Finally, Tead1 −/−; Tead2 −/− embryos showed reduced cell proliferation and increased apoptosis. These results suggest that Tead1 and Tead2 are functionally redundant, use YAP as a major coactivator, and support notochord maintenance as well as cell proliferation and survival in mouse development.

scholarly journals Mice Deficient in the Axonemal Protein Tektin-t Exhibit Male Infertility and Immotile-Cilium Syndrome Due to Impaired Inner Arm Dynein Function

2004 ◽  
Vol 24 (18) ◽  
pp. 7958-7964 ◽  
Author(s):  
Hiromitsu Tanaka ◽  
Naoko Iguchi ◽  
Yoshiro Toyama ◽  
Kouichi Kitamura ◽  
Tohru Takahashi ◽  
...  
Keyword(s):  
Physiological Role ◽  
Electron Microscopic Examination ◽  
Electron Microscopic ◽  
Vitro Fertilization ◽  
Homozygous Mutant ◽  
Causal Genes ◽  
And Function ◽  
Ciliary Dyskinesia

ABSTRACT The haploid germ cell-specific Tektin-t protein is a member of the Tektin family of proteins that form filaments in flagellar, ciliary, and axonemal microtubules. To investigate the physiological role of Tektin-t, we generated mice with a mutation in the tektin-t gene. The homozygous mutant males were infertile, while the females were fully fertile. Sperm morphology and function were abnormal, with frequent bending of the sperm flagella and marked defects in motility. In vitro fertilization assays showed that the defective spermatozoa were able to fertilize eggs. Electron microscopic examination showed that the dynein inner arm structure was disrupted in the sperm flagella of tektin-t-deficient mice. Furthermore, homozygous mutant mice had functionally defective tracheal cilia, as evidenced by altered dynein arm morphology. These results indicate that Tektin-t participates in dynein inner arm formation or attachment and that the loss of Tektin-t results in impaired motility of both flagella and cilia. Therefore, the tektin-t gene is one of the causal genes for immotile-cilium syndrome/primary ciliary dyskinesia.

scholarly journals Smad4 is critical for self-renewal of hematopoietic stem cells

2007 ◽  
Vol 204 (3) ◽  
pp. 467-474 ◽  
Author(s):  
Göran Karlsson ◽  
Ulrika Blank ◽  
Jennifer L. Moody ◽  
Mats Ehinger ◽  
Sofie Singbrant ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
In Vitro Proliferation ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Early Embryonic Lethality

Members of the transforming growth factor β (TGF-β) superfamily of growth factors have been shown to regulate the in vitro proliferation and maintenance of hematopoietic stem cells (HSCs). Working at a common level of convergence for all TGF-β superfamily signals, Smad4 is key in orchestrating these effects. The role of Smad4 in HSC function has remained elusive because of the early embryonic lethality of the conventional knockout. We clarify its role by using an inducible model of Smad4 deletion coupled with transplantation experiments. Remarkably, systemic induction of Smad4 deletion through activation of MxCre was incompatible with survival 4 wk after induction because of anemia and histopathological changes in the colonic mucosa. Isolation of Smad4 deletion to the hematopoietic system via several transplantation approaches demonstrated a role for Smad4 in the maintenance of HSC self-renewal and reconstituting capacity, leaving homing potential, viability, and differentiation intact. Furthermore, the observed down-regulation of notch1 and c-myc in Smad4−/− primitive cells places Smad4 within a network of genes involved in the regulation HSC renewal.

scholarly journals Hypoxia inhibits expression and function of mitochondrial thioredoxin 2 to promote pulmonary hypertension

2017 ◽  
Vol 312 (5) ◽  
pp. L599-L608 ◽  
Author(s):  
Sherry E. Adesina ◽  
Brandy E. Wade ◽  
Kaiser M. Bijli ◽  
Bum-Yong Kang ◽  
Clintoria R. Williams ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Pulmonary Hypertension ◽  
Right Ventricular ◽  
Intracellular Signaling ◽  
Dominant Negative ◽  
Therapeutic Effects ◽  
Thioredoxin 2

Pulmonary hypertension (PH) is characterized by increased pulmonary vascular resistance, pulmonary vascular remodeling, and increased pulmonary vascular pressures that often result in right ventricular dysfunction, leading to right heart failure. Evidence suggests that reactive oxygen species (ROS) contribute to PH pathogenesis by altering pulmonary vascular cell proliferation and intracellular signaling pathways. However, the role of mitochondrial antioxidants and oxidant-derived stress signaling in the development of hypoxia-induced PH is largely unknown. Therefore, we examined the role of the major mitochondrial redox regulator thioredoxin 2 (Trx2). Levels of Trx2 mRNA and protein were examined in human pulmonary arterial endothelial cells (HPAECs) and smooth muscle cells (HPASMCs) exposed to hypoxia, a common stimulus for PH, for 72 h. Hypoxia decreased Trx2 mRNA and protein levels. In vitro overexpression of Trx2 reduced hypoxia-induced H2O2 production. The effects of increased Trx2 protein level were examined in transgenic mice expressing human Trx2 (TghTrx2) that were exposed to hypoxia (10% O2) for 3 wk. TghTrx2 mice exposed to hypoxia had exacerbated increases in right ventricular systolic pressures, right ventricular hypertrophy, and increased ROS in the lung tissue. Trx2 overexpression did not attenuate hypoxia-induced increases in Trx2 oxidation or Nox4 expression. Expression of a dominant negative C93S Trx2 mutant that mimics Trx2 oxidation exacerbated hypoxia-induced increases in HPASMC H2O2 levels and cell proliferation. In conclusion, Trx2 overexpression failed to attenuate hypoxia-induced HPASMC proliferation in vitro or hypoxia-induced PH in vivo. These findings indicate that strategies to enhance Trx2 expression are unlikely to exert therapeutic effects in PH pathogenesis.

scholarly journals Placenta Defects and Embryonic Lethality Resulting from Disruption of Mouse Hydroxysteroid (17-β) Dehydrogenase 2 Gene

2008 ◽  
Vol 22 (3) ◽  
pp. 665-675 ◽  
Author(s):  
Pia Rantakari ◽  
Leena Strauss ◽  
Riku Kiviranta ◽  
Heidi Lagerbohm ◽  
Jenni Paviala ◽  
...  
Keyword(s):  
Embryonic Stem ◽  
Basal Layer ◽  
Embryonic Lethality ◽  
Cellular Density ◽  
Targeted Gene Disruption ◽  
Structural Abnormalities ◽  
The Brain

Abstract Hydroxysteroid (17-β) dehydrogenase 2 (HSD17B2) is a member of aldo-keto reductase superfamily, known to catalyze the inactivation of 17β-hydroxysteroids to less active 17-keto forms and catalyze the conversion of 20α-hydroxyprogesterone to progesterone in vitro. To examine the role of HSD17B2 in vivo, we generated mice deficient in Hsd17b2 [HSD17B2 knockout (KO)] by a targeted gene disruption in embryonic stem cells. From the homozygous mice carrying the disrupted Hsd17b2, 70% showed embryonic lethality appearing at the age of embryonic d 11.5 onward. The embryonic lethality was associated with reduced placental size measured at embryonic d 17.5. The HSD17B2KO mice placentas presented with structural abnormalities in all three major layers: the decidua, spongiotrophoblast, and labyrinth. Most notable was the disruption of the spongiotrophoblast and labyrinthine layers, together with liquid-filled cysts in the junctional region and the basal layer. Treatments with an antiestrogen or progesterone did not rescue the embryonic lethality or the placenta defect in the homozygous mice. In hybrid background used, 24% of HSD17B2KO mice survived through the fetal period but were born growth retarded and displayed a phenotype in the brain with enlargement of ventricles, abnormal laminar organization, and increased cellular density in the cortex. Furthermore, the HSD17B2KO mice had unilateral renal degeneration, the affected kidney frequently appearing as a fluid-filled sac. Our results provide evidence for a role for HSD17B2 enzyme in the cellular organization of the mouse placenta.

scholarly journals The role of NER factors in mouse development and disease

2014 ◽  
Author(s):  
Ισμήνη Καρακασιλιώτη
Keyword(s):  
Mouse Development

Ο ρόλος της γενωμικής αστάθειας στην εμφάνιση ιστο-ειδικής παθολογίας σε ασθενείς και ποντίκια που φέρουν μεταλλαγές στα γονίδια του μηχανισμού εκτομής νουκλεοτιδίων δεν είναι γνωστός και υποθέτουμε πως υπάρχουν ιστο-ειδικές αποκρίσεις κατά επιβλαβών καταστάσεων. Κάνοντας χρήση του Cre/LoxP γενετικού συστήματος και του Fabp4-aP2 υποκινητή που εκφράζεται ειδικά στον λιπώδη ιστό, αναπτύξαμε έναν ποντικό μοντέλο με ιστο-ειδική απώλεια λειτουργίας για το Ercc1 γονίδιο στο λίπος. Τα aP2-Ercc1-/- ποντίκια αναπτύσσονται φυσιολογικά και δεν διαφέρουν από τα ποντίκια αναφοράς. Από την ηλικία των τριών μηνών, τα aP2-Ercc1-/- ποντίκια έχουν λιγότερο βάρος, οι αποθήκες λίπους τους είναι σημαντικά μικρότερες και εμφανίζουν συνολική απώλεια του λιπώδους ιστού. Πειράματα ηλεκτρονικής μικροσκοπίας σάρωσης αποκάλυψαν έναν δραματικό εκφυλιστικό φαινότυπο που περιλαμβάνει ρήξη της βασικής μεμβράνης, απώλεια λιποκυττάρων και εκτενή ίνωση. Ο φαινότυπος αυτός συνοδεύεται από επιπλοκές στον μεταβολισμό που εμφανίζονται συνήθως όταν τα λιποκύτταρα είναι δυσλειτουργικά. Ανάλυση της γονιδιακής έκφρασης του λιπώδους ιστού από αγρίου τύπου και Ercc1-/- ποντίκια κατέδειξε 2200 διαφορικά εκφραζόμενα γονίδια που ανήκουν σε μονοπάτια όπως: απόκριση σε διπλές θραύσεις της DNA έλικας, προ-φλεγμονώδης σηματοδότηση και σηματοδότηση από πυρηνικούς υποδοχείς και αυξητικούς παράγοντες. Μια ομάδα γονιδίων που αποκρίνονται στις διπλές θραύσεις της DNA έλικας υπερεκφράζονται στον λιπώδη ιστό των aP2-Ercc1-/- ποντικιών και κάποια από αυτά συγκεντρώνονται σε πυρηνικές εστίες, η παρουσία των οποίων υποδηλώνει απόκριση σε DNA αλλοιώσεις. Στον λιπώδη ιστό των aP2-Ercc1-/- ποντικιών πολλά λιποκύτταρα πεθαίνουν μέσω νέκρωσης και γύρω τους σχηματίζονται στεφανιαίες δομές, που περιλαμβάνουν προ-φλεγμονώδη λευκοκύτταρα. Η παρατήρηση αυτή σηματοδοτεί την in vivo συσχέτιση της απόκρισης σε DNA αλλοιώσεις με τη φλεγμονώδη απόκριση. Χρησιμοποιώντας ένα in vitro σύστημα κυτταρικών καλλιεργειών, αποδείξαμε ότι η απώλεια της Ercc1, καθώς και οι ίδιες οι DNA αλλοιώσεις, δρουν με κυτταρικά αυτόνομο τρόπο, ώστε να επάγουν την ATM-εξαρτώμενη αντιστροφή της καταστολής υποκινητών που σχετίζονται με την προ-φλεγμονώδη απόκριση, σε ινοβλάστες που έχουν υποστεί διαφοροποίηση προς λιποκύτταρα. Συνολικά, τα δεδομένα αυτά υποστηρίζουν την ύπαρξη ενός ενδογενούς σήματος που εξαρτάται από τις DNA αλλοιώσεις και επάγει μια αυτό-φλεγμονώδη απόκριση στον λιπώδη ιστό, η οποία τελικά οδηγεί στη λιποδυστροφία που παρατηρείται στη NER προγηρία.

scholarly journals Naa12 rescues embryonic lethality in Naa10-Deficient Mice in the amino-terminal acetylation pathway

2020 ◽  
Author(s):  
Hyae Yon Kweon ◽  
Mi-Ni Lee ◽  
Max Dörfel ◽  
Seungwoon Seo ◽  
Leah Gottlieb ◽  
...  
Keyword(s):  
Embryonic Lethality ◽  
Mouse Development ◽  
Post Translational Modifications ◽  
Amino Terminal ◽  
Neonatal Lethality ◽  
Enormous Amount ◽  
Deficient Mice ◽  
Urogenital Anomalies

AbstractThere is an enormous amount of variation in proteins introduced by co- and post-translational modifications, including N-terminal acetylation (NTA), catalyzed by a set of N-terminal acetyltransferases (NATs). The NatA complex (including X-linked Naa10 and Naa15) is the major acetyltransferase, with 40–50% of all mammalian proteins being potential substrates. However, the overall role of NTA on a whole-organism level is poorly understood, particularly in mammals. Male mice lacking Naa10 show no globally apparent in vivo NTA impairment and, surprisingly, do not exhibit embryonic lethality. Rather Naa10 nulls display increased neonatal lethality, and the majority of surviving undersized mutants exhibit a combination of hydrocephaly, cardiac defects, homeotic anterior transformation (including an extra thoracic rib), piebaldism and urogenital anomalies. The lack of complete embryonic lethality in Naa10-null mice is explained by the discovery of Naa12, a previously unannotated Naa10-like paralogue with NAT activity that genetically compensates for Naa10. Mice deficient for Naa12 have no apparent phenotype, except for decreased fertility, whereas mice doubly deficient for Naa10 and Naa12 display embryonic lethality, thus presenting the complete machinery for NatA-mediated NTA in mouse development.

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