Characterization of a Mouse Factor IX cDNA and Developmental Regulation of the Factor IX Gene Expression in Liver

SummaryA mouse factor IX cDNA was isolated and characterrzed. The cDNA was 1,837 bp in length and contained the coding region as well as short 5’ and 3’ untranslated sequences. Northern blot analysis of liver RNA showed two mRNA species of 3.2 kb (major) and 2.2 kb (minor) for the mouse factor IX. An antisgnse RNA probe prepared from the mouse cDNA was employed to determine the steady state level of factor IX mRNA in mouse liver at various developmental stages. The factor IX mRNA level was very low (2–5% of the adult level) during the gestational period until day –3 (gestational day 17) followed by a rapid increase at day –2 through birth. This phase of rapid increase was followed by a gradual increase before it reached the adult level at around 20 to 24 days. At birth, the factor IX mRNA level was found to be at about 43% of that of the adult. The rnRNA levels in mouse liver agreed well with the plasma factor IX activity levels. These results indicate that reduced factor IX activity in newborns is due to the low levels of factor IX mRNA available for translation.

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Age and Sex Dependent Regulation of the Factor IX Gene in Mice

Thrombosis and Haemostasis ◽

10.1055/s-0038-1650693 ◽

1996 ◽

Vol 76 (06) ◽

pp. 0965-0969 ◽

Cited By ~ 12

Author(s):

Sumiko Kurachi ◽

Eri Hitomi ◽

Kotoku Kurachi

Keyword(s):

Reference Group ◽

Mrna Level ◽

Factor Ix ◽

Mouse Strains ◽

Female Group ◽

Human Populations ◽

Activity Levels ◽

Mrna Levels ◽

Animal Groups ◽

Plasma Factor

SummaryPlasma factor IX and liver factor IX mRNA levels in two normal mouse strains (B6D2F1 and BALB/CJNIA) were determined in relation to aging and sex of the animals. With male B6D2F1 mice, mean plasma factor IX activity levels for the 14 and 21-22 month-old animals were found to be 124% and 226%, respectively, of the 5 month-old group. Similarly, liver factor IX mRNA levels for the same age animal groups were 145% and 227%, respectively, of the reference group. Mean plasma factor IX levels for the same age female animals were 132% and 175%, respectively, and were accompanied by similarly elevated liver factor IX mRNA levels, 119 and 175%, respectively, of the 5 month-old female group. Factor IX activity and mRNA levels for the 5,14 and 21-22 month-old female animal groups were lower than those of the corresponding male age groups by 25, 20 and 37%, and 20,36 and 38%, respectively. With BALB/CJNIA mice, similar correlation was observed between the advancing age and substantial elevations in the factor IX mRNA level as well as on the unequal factor IX mRNA levels in females and males.These results indicate that the plasma factor IX level in both male and female mice is greatly elevated with aging, in general agreement with a similar phenomenon observed for human populations, and that this increase is due to a similar elevation in the factor IX mRNA level in the liver. In mice, both factor IX activity and mRNA levels are significantly higher in males than in females, which has not been described for humans.

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Characterization of Genomic Clones and Expression Analysis of the Three Types of Superoxide Dismutases During Nodule Development in Lotus japonicus

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-20-3-0262 ◽

2007 ◽

Vol 20 (3) ◽

pp. 262-275 ◽

Cited By ~ 27

Author(s):

Maria C. Rubio ◽

Manuel Becana ◽

Shusei Sato ◽

Euan K. James ◽

Satoshi Tabata ◽

...

Keyword(s):

Enzyme Activity ◽

Developmental Stages ◽

Expression Patterns ◽

Mrna Level ◽

Lotus Japonicus ◽

Nodule Development ◽

Superoxide Dismutases ◽

Activity Levels ◽

Primary Role ◽

Infection Threads

Superoxide dismutases (SODs) are metalloenzymes that play a primary role in the protection against oxidative stress in plants and other organisms. We have characterized four SOD genes in Lotus japonicus and have analyzed their expression in roots and four developmental stages of nodules. The expression of cytosolic CuZnSOD, at the mRNA, protein, and enzyme activity levels, decreases with nodule age, and the protein is localized in the dividing cells and infection threads of emergent nodules and in the infected cells of young nodules. The mitochondrial MnSOD was downregulated, whereas the bacteroidal MnSOD displayed maximal protein and enzyme activity levels in older nodules. Two additional genes, encoding plastidic (FeSOD1) and cytosolic (FeSOD2) FeSOD isoforms, were identified and mapped. The genes are located in different chromosomes and show differential expression. The FeSOD1 mRNA level did not change during nodule development, whereas FeSOD2 was upregulated. The distinct expression patterns of the SOD genes may reflect different regulatory mechanisms of the enzyme activities during nodule ontogeny. In particular, at the mRNA and activity levels, the virtual loss of cytosolic CuZnSOD in mature and old nodules, concomitant with the induction of FeSOD2, suggests that the two enzymes may functionally compensate each other in the cytosol at the late stages of nodule development.

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Characterization of the Original Christmas Disease Mutation (Cysteine 206 → Serine): From Clinical Recognition to Molecular Pathogenesis

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648381 ◽

1992 ◽

Vol 67 (01) ◽

pp. 063-065 ◽

Cited By ~ 6

Author(s):

Sherryl A M Taylor ◽

Jacalyn Duffin ◽

Cherie Cameron ◽

Jerome Teitel ◽

Bernadette Garvey ◽

...

Keyword(s):

Nucleotide Substitution ◽

Novel Mutation ◽

Factor Ix ◽

Causative Mutation ◽

Coding Region ◽

Body Of Knowledge ◽

Polymerase Chain ◽

Splice Junctions

SummaryChristmas disease was first reported as a distinct clinical entity in two manuscripts published in 1952 (1, 2). The eponym associated with this disorder, is the surname of the first patient examined in detail and reported by Biggs and colleagues in a paper describing the clinical and laboratory features of seven affected individuals (3). This patient has severe factor IX coagulant deficiency (less than 0.01 units/ml) and no detectable circulating factor IX antigen (less than 0.01 units/ml). Coding sequence and splice junctions of the factor IX gene from this patient have been amplified in vitro through the polymerase chain reaction (PCR). One nucleotide substitution was identified at nucleotide 30,070 where a guanine was replaced by a cytosine. This mutation alters the amino acid encoded at position 206 in the factor IX protein from cysteine to serine. The non conservative nature of this substitution, the absence of this change in more than 200 previously sequenced factor IX genes and the fact that the remainder of the coding region of this gene was normal, all provide strong circumstantial evidence in favour of this change being the causative mutation in this patient. The molecular characterization of this novel mutation in the index case of Christmas disease, contributes to the rapidly expanding body of knowledge pertaining to Christmas disease pathogenesis.

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Sequence of the human glycogen-associated regulatory subunit of type 1 protein phosphatase and analysis of its coding region and mRNA level in muscle from patients with NIDDM

Diabetes ◽

10.2337/diabetes.43.10.1234 ◽

1994 ◽

Vol 43 (10) ◽

pp. 1234-1241 ◽

Cited By ~ 26

Author(s):

Y. H. Chen ◽

L. Hansen ◽

M. X. Chen ◽

C. Bjorbaek ◽

H. Vestergaard ◽

...

Keyword(s):

Protein Phosphatase ◽

Mrna Level ◽

Regulatory Subunit ◽

Coding Region

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Yeast CBP1 mRNA 3' end formation is regulated during the induction of mitochondrial function

Molecular and Cellular Biology ◽

10.1128/mcb.11.2.813-821.1991 ◽

1991 ◽

Vol 11 (2) ◽

pp. 813-821

Author(s):

S A Mayer ◽

C L Dieckmann

Keyword(s):

Mitochondrial Function ◽

Developmental Stages ◽

Single Gene ◽

Mitochondrial Protein ◽

Nuclear Gene ◽

State Level ◽

Yeast Cells ◽

Cdna Clones ◽

Coding Sequence ◽

Polyadenylation Sites

Alternative mRNA processing is one mechanism for generating two or more polypeptides from a single gene. While many mammalian genes contain multiple mRNA 3' cleavage and polyadenylation signals that change the coding sequence of the mature mRNA when used at different developmental stages or in different tissues, only one yeast gene has been identified with this capacity. The Saccharomyces cerevisiae nuclear gene CPB1 encodes a mitochondrial protein that is required for cytochrome b mRNA stability. This 66-kDa protein is encoded by a 2.2-kb mRNA transcribed from CPB1. Previously we showed that a second 1.2-kb transcript is initiated at the CBP1 promoter but has a 3' end near the middle of the coding sequence. Furthermore, it was shown that the ratio of the steady-state level of 2.2-kb CBP1 message to 1.2-kb message decreases 10-fold during the induction of mitochondrial function, while the combined levels of both messages remain constant. Having proposed that regulation of 3' end formation dictates the amount of each CBP1 transcript, we now show that a 146-bp fragment from the middle of CBP1 is sufficient to direct carbon source-regulated production of two transcripts when inserted into the yeast URA3 gene. This fragment contains seven polyadenylation sites for the wild-type 1.2-kb mRNA, as mapped by sequence analysis of CBP1 cDNA clones. Deletion mutations upstream of the polyadenylation sites abolished formation of the 1.2-kb transcript, whereas deletion of three of the sites only led to a reduction in abundance of the 1.2-kb mRNA. Our results indicate that regulation of the abundance of both CBP1 transcripts is controlled by elements in a short segment of the gene that directs 3' end formation of the 1.2-kb transcript, a unique case in yeast cells.

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Inherent hepatocytic heterogeneity determines expression and retention of edited F9 alleles post-AAV/CRISPR infusion

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2110887118 ◽

2021 ◽

Vol 118 (42) ◽

pp. e2110887118

Author(s):

Qiang Wang ◽

Lin Zhang ◽

Guo-Wei Zhang ◽

Jian-Hua Mao ◽

Xiao-Dong Xi ◽

...

Keyword(s):

Viral Vectors ◽

Gene Editing ◽

Regulatory Region ◽

Factor Ix ◽

Host Cells ◽

Clotting Factor ◽

Dependent Manner ◽

Coding Region ◽

Therapeutic Benefits

Infusing CRISPR/donor-loaded adeno-associated viral vectors (AAV/CRISPR) could enable in vivo hepatic gene editing to remedy hemophilia B (HB) with inherited deficiency of clotting factor IX (FIX). Yet, current regimens focus on correcting HB with simple mutations in the coding region of the F9, overlooking those carrying complicated mutations involving the regulatory region. Moreover, a possible adverse effect of treatment-related inflammation remains unaddressed. Here we report that a single DNA cutting-mediated long-range replacement restored the FIX-encoding function of a mutant F9 (mF9) carrying both regulatory and coding defects in a severe mouse HB model, wherein incorporation of a synthetic Alb enhancer/promoter-mimic (P2) ensured FIX elevation to clinically meaningful levels. Through single-cell RNA sequencing (scRNA-seq) of liver tissues, we revealed that a subclinical hepatic inflammation post-AAV/CRISPR administration regulated the vulnerability of the edited mF9-harboring host cells to cytotoxic T lymphocytes (CTLs) and the P2 activity in a hepatocytic subset–dependent manner via modulating specific sets of liver-enriched transcription factors (LETFs). Collectively, our study establishes an AAV/CRISPR-mediated gene-editing protocol applicable to complicated monogenetic disorders, underscoring the potentiality of improving therapeutic benefits through managing inflammation.

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Effects of male and female personality on sexual cannibalism in the Springbok mantis

10.32942/osf.io/prq5x ◽

2021 ◽

Author(s):

Pietro Pollo ◽

Nathan W Burke ◽

Gregory I Holwell

Keyword(s):

Personality Traits ◽

Developmental Stages ◽

Male Mating ◽

Sexual Cannibalism ◽

Activity Levels ◽

Male And Female ◽

Young Females ◽

Behavioural Syndromes ◽

Male Activity ◽

Spillover Hypothesis

Behaviours that are consistent across contexts (also known as behavioural syndromes) can have evolutionary implications, but their role in scenarios where the sexes conflict, such as sexual cannibalism, is poorly understood. The aggressive spillover hypothesis proposes that cannibalistic attacks during adulthood may depend on female aggressiveness during earlier developmental stages, but evidence for this hypothesis is scarce. Male activity may also influence sexual cannibalism if males approach females quickly and carelessly, yet this has not been explored. Here we use the Springbok mantis, Miomantis caffra, to explore whether male activity levels and female aggressiveness can explain high rates of sexual cannibalism prior to copulation. We show that male and female personality traits affect male mating decisions, but not sexual cannibalism. Females that were aggressive as juveniles were not more likely to cannibalize males when adult, but these females were approached by males more frequently. More active males were more likely to approach females, but they were neither faster at doing so nor were they more likely to be cannibalized. We also found that size and age influenced mating decisions of both sexes: young females were more like to cannibalize males while young and large males took longer to approach females. Taken together, our results suggest that several traits, including personality, play a role in sexual encounters in M. caffra. Our study further highlights the importance of examining the traits of both sexes when assessing mating dynamics, especially in the context of sexual cannibalism.

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Mammalian nonsense codons can be cis effectors of nuclear mRNA half-life

Molecular and Cellular Biology ◽

10.1128/mcb.14.12.8219-8228.1994 ◽

1994 ◽

Vol 14 (12) ◽

pp. 8219-8228

Author(s):

P Belgrader ◽

J Cheng ◽

X Zhou ◽

L S Stephenson ◽

L E Maquat

Keyword(s):

Triosephosphate Isomerase ◽

Blot Hybridization ◽

Mrna Level ◽

Coding Region ◽

Protein Coding ◽

Codon Recognition ◽

Reverse Transcription Pcr ◽

Nonsense Codon ◽

Bona Fide ◽

Nonsense Codons

Frameshift and nonsense mutations within the gene for human triosephosphate isomerase (TPI) that generate a nonsense codon within the first three-fourths of the protein coding region have been found to reduce the abundance of the product mRNA that copurifies with nuclei. The cellular process and location of the nonsense codon-mediated reduction have proven difficult to elucidate for technical reasons. We show here, using electron microscopy to judge the purity of isolated nuclei, that the previously established reduction to 25% of the normal mRNA level is evident for nuclei that are free of detectable cytoplasmic contamination. Therefore, the reduction is likely to be characteristic of bona fide nuclear RNA. Fully spliced nuclear mRNA is identified by Northern (RNA) blot hybridization and a reverse transcription-PCR assay as the species that undergoes decay in experiments that used the human c-fos promoter to elicit a burst and subsequent shutoff of TPI gene transcription upon the addition of serum to serum-deprived cells. Finally, the finding that deletion of a 5' splice site of the TPI gene results predominantly but not exclusively in the removal by splicing (i.e., skipping) of the upstream exon as a part of the flanking introns has been used to demonstrate that decay is specific to those mRNA products that maintain the nonsense codon. This result, together with our previous results that implicate translation by ribosomes and charged tRNAs in the decay mechanism, indicate that nonsense codon recognition takes place after splicing and triggers decay solely in cis. The possibility that decay takes place during the process of mRNA export from the nucleus to the cytoplasm is discussed.

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Plasma factor IX levels in patients given hexoestrol or stilboestrol to suppress lactation

BMJ ◽

10.1136/bmj.4.5675.82 ◽

1969 ◽

Vol 4 (5675) ◽

pp. 82-84 ◽

Cited By ~ 14

Author(s):

C. A. Hakim ◽

M. G. Elder ◽

D. F. Hawkins

Keyword(s):

Factor Ix ◽

Plasma Factor

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Further Experience on the Effect of High Pre-Donation Factor VIII Levels in Blood Donors on the Factor VIII Content of Small-Pool Fractions

10.1055/s-0039-1682568 ◽

1977 ◽

Author(s):

H. Beeser ◽

H. Eqli

Keyword(s):

Factor Viii ◽

High Plasma ◽

Activity Levels ◽

Factor Viii Activity ◽

Factor Viii Level ◽

Plasma Factor ◽

Viii Level ◽

Factor Viii Concentrates ◽

Small Pool ◽

Concentrate Preparation

Because of the well known wide normal range of the factor VIII activity between 60 to 170% I man, selecting of donors with high activity levels would be of advantaae for the preparation of factor VIII concentrates. This is especially true for preparing small-pool fractions, as for technical reasons the final product cannot be controlled for its factor VIII content. In preliminary investigations, we reported on elsewhere, high factor VIII activity in donors estimated before a donation had been rarely reproducible before a second donation after 8-12 weeks. So as a preliminary result of finding a donor’s factor VIII level varying from donation to donation selecting of plasmas with high factor VIII content for concentrate preparation could only be establishedby re-estimating the activity before each donation. Proceeding in this way would be much too troublesome. To get more reliable information whether a healthy subject’s high factor VIII plasma level is distinctly varying or rather constant we assayed the plasma of 200 donors with factor VIII activity > 120% two times more before donation. The results confirmed our preliminary findings, especially the fact that a high plasma factor VIII activity in experienced donors was rarely reproducible when re-estimated before a second and third donation. As a consequence selecting of donors with high factor VIII procoaqulant activity for preparing small-pool factor VIII concentrates is impracticable.

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