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Type I Factor XI11 Deficiency Is Caused By a Genetic Defect of Its b
Subunit: Insertion of Triplet AAC in Exon I11 Leads to Premature
Termination in the Second Sushi Domain
By Tornonori Izurni, Teruto Hashiguchi, Giancarlo Castaman, Alberto Tosetto, Francesco Rodeghiero,
Antonio Girolarni, and Akitada lchinose
Factor Xlll deficiency has been classified into two categories:
type I deficiency, characterized by the lack of both thea and
b subunits; and type II deficiency, characterized by the lack
of the a subunit alone. To clarify the genetic bases of these
diseases, previously reported cases of the type I deficiency
were examined at the DNA level. DNA sequence analysis
showed that a nucleotide triplet (AAC) was inserted within
the codon for Tyr-80 in exon 111 of the gene for a female
proband‘s b subunit, resulting in the creation of a stop codon. Restriction digestion of amplified DNAs confirmed that
the proband and hersister were homozygotes, and their
family members were heterozygotes of this mutant allele.
A truncated protein composed of 79 amino acids could be
synthesized by these homozygotes; however, such aprotein
would not besecreted or it would degrade quickly, because
there were normal amounts of the mutant mRNA, but no b
subunit was detected in these patients. The a subunit deficiency of these patients must be secondary to theb subunit
deficiency, as their gene for the a subunit was intact, and
the a subunit in their platelets was present within normal
levels.
0 1996 by The American Societyof Hematology.
F
a change of Cys430 to Phe in the seventh Sushi domain
causes impaired transport of the mutant b subunit, resulting
in its deficiency.”
In this study, to clarify the genetic basis of type I factor
XI11 deficiency, we examined the two original patients6.*’ at
the DNA level and found that they are homozygotes of a
defective b subunit allele. Accordingly, a new genetic classification for factor XI11 deficiency is proposed.
ACTOR XI11 deficiency is an inherited hemorrhagic disease characterized by a life-long bleeding tendency and
abnormal wound healing in affected patients and spontaneous abortion in affected fernales.l~*Coagulation factor XI11
is a plasma transglutaminase consisting of two catalytic a
and two noncatalytic b s~bunits.’.~.~
The a subunit contains
an active site region of transglutaminases, and the b subunit
is composed of 10 tandem repeats called “Sushi domain^"^
because of their shape, or “short consensus repeats (SCRs)”,
which are also called “GP-l structures” because they were
identified first in P*-glycoprotein I. Factor XI11 activated
by thrombin catalyzes intermolecular cross-linking reactions
between various proteins in plasma and an extracellular matrix. Thus, deficiency of factor XI11 results in defective crosslinking reactions.
More than 200 cases of factor XI11 deficiency have been
identified. Its frequency is calculated to be one in 10,000.*
In most cases, the diagnosis for factor XI11 deficiency is
made by the measurement of factor XI11 activity, which
represents the amount of the functional a subunit. It had
been proposed by Girolami et aP7that factor XI11 deficiency
is to be classified by the presence or absence of antigens
into two groups: type I deficiency, characterized by the lack
of both a and b subunits, and type I1 by the lack of a subunit
alone. This classification canbemadeonlyby
a precise
immunologic examination employing monospecific antibodies against the a and b subunits, separately. Unfortunately,
an exact classification for factor XI11 deficiency has been
difficult and confusing because at least one commercial antiserum against the b subunit crossreacted with another plasma
protein as reported in 19786and several vials of commercial
antibodies have done so during the last 5 years in our laboratories. Thus, some of the patients of real type I deficiency
could have been misdiagnosed as having type I1 deficiency
by erroneous immunologic measurements.
Determination of the genomic sequence for both subunits’”’
has made it possible to characterize factor XI11 deficiency
decisively at the DNA level.‘*The several examples of type
II factor XIII deficiencyinwhichthemutationis
knownl*-“j
all result from genetic defects of the a subunit. An inherited
quantitative abnormality of the b subunit also leads to factor
XI11 deficiency,17in which two genetic mutations in the gene
for the b subunit have now been identified.” In this patient,
Blood, Vol 87, No 7 (April l ) , 1996: pp 2769-2774
MATERIALSANDMETHODS
Study subjects. Venous blood was drawn from normal individuals, two previously reported patients with type I factor XIII deficiency‘,*” and their family members (Fig 1) after informed consent
had been obtained. Factor XI11 activity was determined by the standard dansylcadaverine-incorporation assay?’ and the a and b subunits in plasma were measured by enzyme-linked immunosorbent
assay (ELISA)?~
In vitro amplification ofthe b subunit gene. Genomic DNA
samples were prepared from leukocytes by standard techniques. A
total of 1 pg of a genomic DNA was amplified in a 100-pL reaction
mixture employing 2.5 to 5.0 U of Thermus aquaticus DNA polymerase (Promega, Madison, W1 or AGS GmbH, Heidelberg, Germany). After 30 cycles of amplification, 9 pL of each reaction mixture was applied to a 0.8% agarose gel containing ethidium bromide
From the Department of Molecular Patho-Biochemistry, Yamagata University School of Medicine, Yamagata, Japan; the Department of Haematology, San Bortolo Hospital, Vicenza, Italy; and the
Institute of Medical Semeiotics, University of Padua Medical School,
Padua, Italy.
Submitted May 22, 1995; accepted November 13, 1995.
Supported in part by research grants from the Mochida Memorial
Foundation (Japan), the Ichiro Kanehara Foundation (Japan), and
the Japan Research Foundation for Clinical Pharmacology.
Presented in part at the 36th Annual Meeting of the American
Society of Hematology, Nashville, TN, December 2-6, 1994.
Address reprint requests to Akitada Ichinose, MD, PhD, Department of Molecular Patho-Biochemistry, Yamagata University School
of Medicine, Iida-Nishi 2-2-2, Yamagata, 990-23 Japan.
The publication costs of this article were defrayed in part by page
charge payment. This article must therefore be hereby marked
“advertisement” in accordance with 18 U.S.C. section 1734 solely to
indicate this fact.
0 1996 by The American Society of Hematology.
0006-4971/96/8707-0$3.00/0
2769
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IZUMI ET AL
2770
(0.5 pg/mL) in 1 X TBE buffer (89 mmol/L Tris base189 m m o K
boric acid/:! m m o K EDTA, pH 7.8). Oligonucleotides were prepared with an Applied Biosystems’ synthesizer. For amplification,
a total of six pairs of gene-specific primers were designed from the
nucleotide sequence of the normal 6 subunit gene” as listed in Table
1 of reference 18 with a slight modification; pair 2R for Exons I1
and 111, S’-side, the sense primer of former pair 3’’ and 5”TCAGGATCCTGCATTGTAGACATAATGAAAAATAA-3’ (antisense);
pair 3R for Exons 111, IV and V, 5”GATGGATCCAAAATGAAATCGCCAATAATAACATT-3’ (sense) and the antisense primer of
former pair 3.”
Nucleotide sequence analysis. The amplified DNA samples were
digested with restriction enzymes to generate the proper ends for
ligation into sequencing vectors. The digested samples were applied
to a 0.8% agarose gel, electroeluted, and then subcloned into
M13mp18 or M13mp19 (GIBCO-BRL, Gathersburg, MD or Toyobo, Tokyo, Japan) with restriction sites to obtain discrete sequences.
The DNA sequence of an insert was then obtained using the dideoxynucleotide method” with deoxyadenosine S’-[a-[’sS]thio]triphosphate (Amersham, Arlington Heights, IL). Tominimize the possibilityof obtaining the DNA sequences with misincorporation by the
Taq DNA polymerase, 10 or more samples of each amplified region
of the b subunit gene were examined.
Detection of the insertion mutation by restriction digestion. For
restriction enzyme analysis for the insertion of a triplet AAC in exon
111, 15 pL of the DNA sample amplified by employing the 5”primer
(sense) of pair 3R and the 3”primer (antisense) of pair 2R was
incubated with 2 U of Mae111 endonuclease (Boehringer Mannheim
GmbH, Germany) for 2 hours at 45°C. The amplified DNA also was
incubated with 1.S U of Tru91 endonuclease (Boehringer, Mannheim,
Germany) for 2 hours at 65°C. After digestion, half of each sample
was applied to a 2% agarose gel.
Immuno6lotting of the b subunit. For Western blotting of the 6
subunit, plasma was mixed with sample buffer (125 mmoUL TrisHCI, pH 6.8, 4% sodium dodecyl sulfate [SDS], SO% glycerol, and
1 mg/mL bromophenol blue) and deionized water at a ratio of 1:5:4,
and then heat-denatured. Samples were electrophoresed on a 10%
polyacrylamide gel. The separated proteins were transferred electrophoretically onto a nitrocellulose membrane, and the membrane was
blocked at 4°C overnight with the blocking buffer containing 0.5%
nonfat dry milk and a washing buffer (1 0 mmol/L Tris-HCI, pH 7.4,
and 150 mmolk NaCI). The blocked membrane was incubated at
room temperature for 1 hour with rabbit antiserum against the 6
subunit (Calbiochem, La Jolla, CA), rinsed three times, and incubated at room temperature with a goat antirabbit IgG (H + L) conjugated with horseradish peroxidase (Bio-Rad, Richmond, CA). The
band for the b subunit was detected by adding diaminobenzidine
and H202.
Reverse transcription-polymerase chain reaction (RT-PCR) assay
of’ aberrant mRNA for the b subunit. Total mRNA was extracted
by the standard guanidinium thiocyanate method from leukocytes
followed by ultracentrifugation on a cesium chloride gradient. The
isolated RNA was stored at -80°C until use. RT of the total RNA
( I .7 pg) was performed using an oligo dT (dT,,) primer and Superscript 11 RNase H- (GIBCO-BRL, Gathersburg, MD). A total of l /
SO of the synthesized first-strand cDNA was used for PCRin a
reaction mixture of 50 pL by employing 0.5 yL of a-[’*P]-dCTP
( 1 0 mCi/mL, ICN Biomedicals Inc, Costa Mesa, CA) and two pairs
of primers separately; for the b subunit, 5”ATCTGTGCAGTGCAACAGAGG-3‘ (sense) and S’-CATTGAATTCTATGTTCTTAAGGG’ITC’ITGATAA-3’ (antisense, the underlined nucleotides were
introduced to create an EcoRI site); for human glyceraldehyde-3phosphate dehydrogenase (GAPDH) as an internal control, 5’-CATCACCATCTTCCAGGAGC-3’ (sense) and S’-TAAGCAGTTGGTGGTGCAGG-3’ (antisense). After 20 cycles (for GAPDH) or 25
cycles (for the 6 subunit), 8 p L of the reaction mixture (GAPDH)
or 15 pL (the 6 subunit) were applied to a 2.5% agarose gel. After
electrophoresis, the gel was dried and subjected to quantitation of
the amplified products by BAS1000 Mac Bio-Imaging Analyzer
(FUJIX, Tokyo, Japan). The fluorescence intensity of the PCR product for the 6 subunit was normalized to that for GAPDH. The dried
gel was also exposed to Kodak XAR autoradiography film (Eastman
Kodak CO, Rochester, NY) at -80°C.
Exarninution of the a subunit. All regions, including exons and
exonhtron boundaries of the a subunit gene, were amplified by
employing 16 pairs of primers as described.” Thesingle-strand conformation polymorphism (SSCP) analyses of the amplified DNAs
were performed as des~ribed.*~.*~
At10°C , a SSCP gelwasrun in
the buffer cooled by Model 1000 Mini Chiller (Bio-Rad). Nucleotide
sequences of the amplified DNA samples were also determined as
described above.
To quantitate the a subunit in platelets by ELBA,’? platelets were
isolated from citrated blood by centrifugation on Ficoll-Hypaque
gradients andwashedin
Tris-EDTA-saline buffer.’” The platelets
were lysed by incubation with 1/40 volume of 20% wt/wt Triton X100.
RESULTS
Insertion of U triplet AAC i n exon III of the gene , f . r the
b subunit. Six pairs of specific amplification primers were
employed for in vitro amplification of all 12 exons andtheir
boundaries of the gene for theb subunit (Fig 2) in the DNA
of theproband’ssister (11-1 in Fig 1). Pairs 1-6 produced
2.1, 1 S , 1.8, 2.0, 2.3, and 1.7 kb DNA fragments, respectively (data not shown). The sizes of these fragments were
exactly the same as predicted from the normal genomic sequence.” These resultsconfirmedthat there was no gross
change(s) in the patient’s gene for the b subunit.
Nucleotide sequences of the six amplified fragments were
then examined to find any possible small difference(s) from
the normalsequence. When exon I11 of the probandwas
sequenced, all of the 15 ssDNA samples showed ATGGTTA U C A T C (Fig 3), while the normal sequence is ATGGTTACATC. The insertion of this AAC triplet falls within
a codon forTyr-80: between its first and secondnucleotides,
-adenosine-5285 and cytidine-5286,” resulting in the creation of a stop codon at amino acid position 80.
Another mutation, adenosine-27970to guanosine, was
also found in the 3’-noncoding region. This is one of the
same polymorphisms that was found in the Japanese patient
with complete h subunit deficiency and also in normal individuals.’* No other change
in the nucleotide sequence of the
proband’s gene for the b subunit was detected by sequence
analysis after the remaining exons andtheir flanking regions
were compared with thenormal gene.“ Thus, it is concluded
that the only genetic defect regarding this patient’s b subunit
is the insertion of the triplet in exon I11 of the gene.
Detection of the insertion by restriction enzyme digestion.
Because insertion of the nucleotide triplet (AAC) in the proband’s DNA destroys an intrinsic Mae111 site (GTNAC) in
exon I11 and creates a new Tru9I site (TTAA), a fragment
spanning the entire region of exon 111 was amplifiedand
digestedwith Mae111 or Tru9I endonuclease. The MueIII
digest of the 430-bp fragment from oneof the family members (111-5) and from a normal individual yielded two fragments of 285 bp and 145 bp (Fig 4, top), whereas the 433-
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2771
GENETIC BASIS OF TYPE I FXlll DEFICIENCY
G
I
A
N123456
T
C
/
N123456
/I
I
T
I1
A
111
50, 54/52
90,54159
58, 60155
50. 50/47
74. 81/55
121, 103/106
Fig 1. Family pedigree of typeI factor Xlll deficiency. Closed circles
represent homozygotes, and half-closed circles (females1and squares
(males) stand for heterozygotes. An arrow indicates the proband
(112); her father (1-1) died before the present study. The first, second,
and third numbers indicate the
b subunit antigen, the activityof the B
subunit, and its antigen levels in plasma, respectively, in each family
member.
bp fragments of the DNAs from the proband and her sister
(11-2 and 11-1) remained unchanged. The remaining five of
their family members (1-2.11-3.111-1,111-2. and 111-3) showed
both cleaved and uncleaved bands. These results indicated
that the two patients of type I deficiency were homozygotes
and five of their family members were heterozygotes of this
mutant allele, which is not found in normal individuals.
The Tr~r91digest of the 430-bp fragments from a normal
individual and a member (111-5) (Fig 4, bottom)yielded five
fragments of 293 bp. 47 bp, 44 bp, 41 bp, and 5 bp (the last
fourfragmentsare
unresolved because ofsmall size). In
contrast, the digest of the 433-bp fragment fromthe proband
and her sister yielded six fragments of 242 bp, 54 bp, 47
bp, 44 bp, 41 bp, and 5 bp(the last five fragmentsare
unresolved). Theremaining five oftheirfamily
members
showed both 293-bp and 242-bp bands. Thus, the genotype
of each individual was confirmed by digesting the fragment
with both endonucleases. This is consistent with the phenotype determined by ELISA (Fig l ) .
Search for the truncated b subunit in plasma. The insertion ofthe AAC triplet falls within a codonforTyr-80,
resulting in the creation of a stop codon at amino acid posi-
.
Sushi Domain
519~1
Gm€
frapmanla
8
5 6 7
I
1-
VI
I1 I V
I I I V
PR-
4-
m-
*
Vlll
VI1
IX
5-
and samples 1 through 6 are from the proband (11-21. Sequences of
all samples were thesame as those of normal except for an insertion
of AAC, shown in a square.
tion 80 (Fig S. top). This would lead to the formation of a
peptide consisting of only79aminoacids
andincluding
only a single Sushi domain (Fig 5, bottom). To investigate
whether thetruncatedhsubunit
is present in circulation,
plasma obtained from two patients of type I deficiency (11-1
and 11-2) and their family members was analyzed by Western
blotting using anti-h subunit antiserum. A band for the normal h subunit of about 70 kD was observed in normal individuals and seven family members. but not in the patients
(data not shown); or were bands for the truncated h subunit
patients'
including
possible
oligomers detected in the
plasma. It is very likely that the resulting truncated protein
does not reach circulation, as no h subunit antigen was detected by more sensitive ELISA in either the proband or her
sister."
Semiquantitation of mRNA for the bsubunit.Because
noprotein for themutant b subunit was detected in the
Mae 111
digest
C-lermlnal
9
:::
c-)"-HH
Exons
PCR
1 2 3 4
Fig 3. Nucleotide sequence of part of exon 111. The left-end lanes
(NI in nucleotide G, A, T, C groups are from the normal individual,
IO
II
LI'NC
m m
l , ,
I
x
X I XI1
e0
For diammair U
Fig 2. Strategy for in vitro amplification of the
gene for the b
subunit and the insertiondetected. All 12 exons and theirboundaries
in the gene for the b subunit were amplified employingsix pairs of
oligonucleotide primers shown byarrows. For analysis of the insertion of AAC between adenosine-5285 and cytidine-5286 shown by a
closed circle, a region was amplified employing the
5"primer of pair
3R and the 3"primer of pair 2R. One of genetic polymorphisms is
shown by an open circle.
100
digest
Fig 4. (Top) Restriction digestionwith Madllendonuclease of the
amplified products of exon 111. (Bottom) Restriction digestion with
TrusI endonuclease. Molecular weight standards are from Bethesda
Research Laboratories. Samples 1-2, 11-1, 11-2, 11-3, 111-1. 111-2, 111-3, and
111-5 refer t o the proband's mother, the probands elder sister, the
proband, her younger sister, et al. as indicated in Fig 1.
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IZUMI ET AL
2772
Exon 111
Normal
1.....77 78 79 8081
82 83......641
H
G
Y
l
S
D .,.... T STP
G A A . . . . A G T ART GGT TRC ATCTCT
GAT..
ACA TAG
E ..... S
....
T
AAC
Mutant
1.....77 78 79
H
G STP
S
GAR
RGT ART GGT TA
E.....
....
ATC TCT G A T . . . . . . .
...
was detected by agarose gel electrophoresis (data not
shown); neither was any mobility shift foundby SSCP analyses of the amplified DNAs when compared with normal
(data
not shown). In addition, no nucleotide substitutions were
detected by sequencing analysis of the amplified DNAsafter
they were compared with the normal genomic sequence of
the n subunit.“’ Furthermore, normal amounts of the n subunit were detected by ELlSA in platelets obtained from the
members of this family (Table I), whereas two patients with
type I1 deficiency from two other independent families had
no n subunit in their platelets. Thus, it was concluded that
the gene for the CI subunit is intact in all members of the
family examined in the present study.
DISCUSSION
The only significant mutation detected in this patient’s h
subunit was the insertion of thetriplet in exon 111 of the
1st Sushi Domain
L
1
II
Xlllb
300
tlmc
T
STP
Fig 5. (Top) Amino acid sequence of the normal and mutant b
subunits. Amino acid sequence of a part of Exon 111 is shownin detail.
Amino acid residues are indicated by a one-letter code. STPstands for
an in-frame stop
codon, created by the mutation. (Bottom)
Schematic
representation of the mutant b subunit. A truncated protein composed of only 79 amino acids would make up a single (first) Sushi
domain, while oneCys residue (with -?l would remainunpaired. The
signal peptide region has been enclosed. Arrows indicate positions
of introns A and B.
patients’ plasma, its mRNA was examined. A RT-PCR assay
was performed to estimate the amount of aberrantly expressed mRNA for the h subunit in leukocytes, as samples
of hepatocytes from these patients were not available. Both
a 493-bp fragment for the h subunit and a 253-bp fragment
for GAPDH were obtained by RT-PCR for a normal Japanese, a homozygote (11-l), and a normal Italian individual
(Fig 6, top). Under experimental conditions, a linear relationship was observed between the amounts of total RNA employed and the fluorescence intensity of these bands (Fig 6,
bottom). The ratio of the h subunit mRNA/GAPDH mRNA
was 0.7, 0.6, and 0.7 for the normal Japanese, the homozygote (11-l), and the normalItalian. respectively. Thus, the
amount of the patient’s mRNA was comparable to that of
normal individuals, strongly suggesting that the mutant
mRNA is synthesized as stable as normal.
Examination of the a subunit. All 15 exons of the gene
for the n subunit of the proband and her sister were successfully amplified by PCR, and no size difference from normal
3ooJ
GAPDH
200
U,
U
0
10
20
30
40
50
Total RNA (ng)
Fig 6. (Top) RT-PCR assay for the mRNAs of the b subunit (Xlllb)
and GAPDH. A radioactive nucleotide was incorporated into
PCR
products t o estimate the amounts of mRNAs for both genes. Four
left of the panel contain amplified
products of
lanes (Control) on the
diluted (1 t o 1/81 samples from a normal individual. Normal4 and
-I stand for samples from normal Japanese and Italian individuals,
respectively, and 11-1 for a homozygote of the typeI deficiency. (Bottom) Linear relationship between the amounts
of total RNA used and
the fluorescence intensity of the radioactive bands. Amounts of the
amplified product roughlyrepresent the amounts of mRNAs for the
b subunit (0)
and for GAPDH (01,respectively.
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2773
GENETIC BASIS OF TYPE I FXlll DEFICIENCY
Table 1. Amounts of a Subunit in Platelets
a Subunit
Subject
Type I deficiency
1-2
11-3
55 1g/i09 platelets
65
58
66
111-1
49
11-1
11-2
111-3
Type II deficiency
60
Case 1
Case 2
Normalrange
Average (n = 10)
Absent*
Absent*
34-78
50
* Lower than detection limit.
gene. Both the proband and her sister were homozygotes,
and five of their family members were heterozygotes of this
mutant allele. Neither normal nor truncated b subunit was
detected in the homozygotes’ plasma. These results suggest
either that the aberrant b subunit is not synthesized or that
it quickly degrades. The mutant mRNAmaybe unstable;
however, it is notpossible to study the effect of this mutation
on the in vivo mRNA for the b subunit because it is synthesizedin the liver and mRNAs of hepatocytes cannot be
obtained from these patients. As an alternative, the amount
of aberrantly expressed mRNA in leukocytes was measured
by a RT-PCR assay. The amount of the patient’s mRNA
was comparable to that of normal individuals, suggesting
that transcription of the gene for the b subunit and stability
of its mRNA are not affected by the mutation. The truncated
peptide contains Cys-71, butnot its counterpart Cys-126,
which would be expected based on its homology with p2glycoprotein I’7 and C4 binding protein.’* Therefore, it is
very likely that this Cys residue in the second Sushi domain
remains unpaired (Fig 5, bottom), whichmay render the
mutant peptide unstable and/or decrease its biosynthesis or
secretion from hepatocytes. A number of cases where deficiencies of various proteins are due to the appearance of
unpaired Cys residues support this conclusion.19~29~3n
In contrast to the genetic defect of the patients’ b subunit,
the genes for the a subunit were intact and expressed in
normal amounts. Accordingly, it is concluded that the a
subunit deficiency of these patients is secondary to the b
subunit deficiency; that is, the b subunit deficiency is the
genetic basis for type I factor XI11 deficiency.
Before the first case of complete b subunit deficiency was
found in Japan,” at least four patients with type I factor XI11
deficiency, demonstrating similar laboratory findings, had
been reported.6,3’,3z
The present study clarified that two of
them were cases of the b subunit deficiency, but the remaining two cases have not been examined for their genetic
defects. They may have separate defects in both the a and
b genes by chance, resulting in real combined deficiency of
both subunits. Alternatively, the lack of the a subunit in
plasma in these cases maybe secondary to the b subunit
deficiency, in the same manner as the Japanese
In
the past, only the enzymatic activity of factor XI11 could be
used for diagnosis of its deficiency, then immunological
assays of each antigen became available, and finally genetic
analyses for both subunits have been developed for precise
diagnosis. When patients with the a subunit deficiency who
are diagnosed by the factor XI11 enzymatic activity are examined, the amount of the b subunit antigen must be determined
because the a subunit deficiency can be caused by genetic
defects either of the a or b subunit. Measurements of the a
subunit in platelets (or placenta, if available) are also useful’7
because, if there is no a subunit in platelets, it may be concluded that the a subunit deficiency is independent of the b
subunit deficiency. Moreover, if normal amounts of the a
subunit are present in platelets, it may be concluded that the
gene for the a subunit is normal and that the gene for the b
subunit must have some defect(s).
The present study showed that the two original patients of
type I deficiency are the second and third cases of complete b
subunit deficiency to be reported thus far. In addition, the
first case with complete b subunit deficiency demonstrated
the a subunit deficiency as well,17 indicating that this patient
falls into the category of type I deficiency. Thus, we concluded that congenital deficiency of the b subunit is the
genetic basis of type I factor XI11 deficiency. On the other
hand, many patients of type I1 factor XI11 deficiency inwhich
the mutation is
all result from genetic defects of
the a subunit. It is, therefore, quite reasonable to conclude
that congenital deficiency of the a subunit is the genetic
basis of type I1 factor XI11 deficiency. There is, as yet, no
example containing real combined genetic defects for both
the a and b subunits. Such a patient would show the same
phenotype as for type I deficiency. Although the combined
genetic deficiency of both subunits could occur, it would be
extremely rare because these genes are localized to separate
chromosomes.” Therefore, we propose a new genetic classification for factor XI11 deficiency in threeparts: the a subunit
deficiency (former type I1 deficiency), the b subunit deficiency (former type I deficiency), and a possible combined
deficiency of both the a and b subunits.
ACKNOWLEDGMENT
The authors thank DrsM. Togashi and S. Tsutsumi for performing
the PCR-SSCP analysis for the a subunit gene and L. Boba for her
help in preparation of the manuscript.
REFERENCES
1 . Lorand L, Losowsky MS, Miloszewski KJM:Human factor
XIII. Fibrin-stabilizing factor. Prog Thromb Hemostas 5:245, 1980
2. Duckert F: Documentation of the plasma factor XI11 deficiency
in man. Ann N Y Acad Sci 202:190, 1972
3. Folk JE, Chung SI: Blood coagulation factor XIII: Relationship
of some biological properties to subunit structure,in Reich E, Rifken
DB, Shaw E (eds): Proteases and Biological Control. New York,
NY, Cold SpringHarborLaboratory, 1975, p 157
4. Ichinose A: Physiology and biochemistry of factor XIII, in
Bloom AL, Forbes CD, Thomas DP, Tuddenham EGD (eds): Haemostasis and Thrombosis. Edinburgh, UK, Churchill Livingstone,
1995, p S31
S. Ichinose A, Bottenus RE, Davie EW: Structure of transglutaminases. J Biol Chem 265:13411, 1990
6. Girolami A, Burul A, Fabris F, Cappellato G, Betterle C: Stud-
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2774
IZUMI ET AL
ies on factor XI11 antigen in congenital factor
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20. Rodeghiero F, Tosetto A, Di Bona E, Castaman G: Clinical
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tic and genetic studies on fibrin-stabilizing factor with a new aswy
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based on amine incorporation. J Clin Invest 48: 1054. I969
acid sequence of the a subunit of human factor XIII. Biochemistry
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22.TosettoA,CastamanG.Rodeghiero
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199s
From www.bloodjournal.org by guest on June 18, 2017. For personal use only.
1996 87: 2769-2774
Type I factor XIII deficiency is caused by a genetic defect of its b
subunit: insertion of triplet AAC in exon III leads to premature
termination in the second Sushi domain
T Izumi, T Hashiguchi, G Castaman, A Tosetto, F Rodeghiero, A Girolami and A Ichinose
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