Diabetes in Human Populations |
Defining Human Populations |
Ethnicity and Population Genetics |
Measuring Human Genetic Polymorphism |
This document was developed by NMSU staff from a presentation made by Drs. Paul Kraemer and P. Scott White, Los Alamos National Laboratory, August 1997 at the workshop "The Human Genome Project: Impact on the Prevalence of Diabetes".
"It has often been said that 'we don't inherit genes, we inherit chromosomes,'" said Dr. Kraemer. Researchers hope that by identifying certain genetic markers with certain ethnic groups, they will gain an insight into the genetic basis of disease and why certain conditions are more prevalent in certain populations. Many scientists believe that you can't change a single base pair anywhere in the genome without having some influence on the body.
There is a higher incidence of diabetes among Native Americans (especially Native Americans from the Southwest) and Hispanic Americans than among the population in general. Whenever a particular group of people shows a predilection towards any disease, a genetic basis for the disease is suspected. [Yearbook of Physical Anthropology 27:153-178 (1984)]
Diabetes has a genetic component, but not one that, on the molecular level, is obvious or well-defined. When talking about a complex disease like diabetes, researchers have to bring in ethnology and social issues. The study of diabetes can't be limited to the study of molecules - scientists must also investigate populations. One ethnic group that has been the focus of diabetes studies is the Native American Pima tribe of Arizona. [American Journal of Human Genetics 57:160-170 (1995)]
Dr. Paul Kraemer of the Los Alamos National Laboratory said
"Most dictionaries define ethnicity as a sizable group of people sharing a common racial, national, religious, linguistic, or cultural heritage. But, there are many components to the word ethnicity that are poorly resolved."
He feels that researchers should try to resolve some of the components of this word, especially the genetic component, because it's the genetic component that may actually give them an avenue for doing some constructive things.
"It's important to remember that there isn't a careful, or a tandem transference of all these components, especially the genetic component. You can have a group of people, and the Pima Indians are a good example of an ethnic group that is clearly defined in cultural terms. It does not necessarily mean that members of this group have retained the genetic component that is Pima."
One way in which people make sense of the world is by categorizing: we look
at a person, and then say something about them. While this human observational
system may be good at identifying people by appearance and behavior, it
doesn't always allow us to identify one another's genetic makeup.
The first part of ethnicity is how people identify and categorize themselves;
the second part is how they are identified and categorized by other people.
One main factor influencing ethnicity is change. Many researchers suspect
that most government statistics on Native Americans are far from accurate.
While numerous Native Americans have retained some ethnic identity in customs
and language, the genetics are changing all the time. Many Native Americans
leave their tribal homes to live in cities where there is a lot of intermingling
of genes through marriage. There are really no accurate records as to how
many people have Native American blood or what percentage it is. This holds
true for Pimas, who have a long history of interaction with other tribes
and other cultures.
One way that researchers follow a genetic line is by studying the language
of a people. Although there are a great many languages spoken in the Americas,
linguistic evolution happens quickly (the Romans spoke only Latin, but just
1,000 years later, all of the romance languages had been developed). By
comparing all of the Native American languages, anthropologists Joseph Greenberg
and Merritt Ruhlen determined that peoples of the new world can be divided
into three groups; Amarinds (all of those living in South and most of North
America); Athabascan, or Na-Dine (Navajos, Apaches and many northern U.S.
and southern Canadian Indians); and Aleut (the Eskimos of the extreme north).
These three groups seem to represent three separate migrations from Asia
into the Americas. The Aleut people have a very low incidence of diabetes,
while in all other Native American peoples, the incidence is high. [Scientific American 267(5): 94-99 (1992)]
Even after the formation of separate tribes and distinct languages, Indian
peoples continued mixing with one another. They took hostages during warfare;
refugees from one area would join a different tribe in another area; and
babies were stolen. Of course, this intermingling among people meant the
blending of genes as well.
One method of tracing ancestry is with the use of surnames. Often, there
is a close vertical transmission of surnames, but there are important exceptions
to that. There are about 20 unique New Mexican names (e.g. Archuleta) that
are not found in Mexico City or Spain. In the new world, many European settlers
took Native American women for wives. It seems that the reason for a higher
incidence of diabetes in Hispanic populations of the Americas is from mixed
marriages with Native Americans.
Ethnographers study a number of polymorphisms, including protein differences,and
various nuclear DNA (using RFLP's, etc.). They have focused heavily on mitochondrial
DNA (mtDNA) for ethnic studies. The mitochondria is a prokaryote derivative
with only a .3% difference among humans around the world.
Some Southwestern Indian tribes have a creation myth of all life emerging
from the Sipapu - a hole in the ground leading up from earlier worlds. There
is a notion of a scientific Sipapu - that is, all life sprung from the same
source and is related by recognizable gene sequences.
Genetic data can be used to tell how closely related a specific individual
is to a specific population. Scientists use a number of methods to compare
genetic material.
One important type of genetic data is chromosome difference, including the number and shapes of chromosomes, and their banding pattern differences.Bands show evolutionary history on a large scale. These bands indicate how two chromosomes fused together, or how one broke, then moved around and picked up somewhere else. By studying fusions and breaks, researchers can "tease apart" some evolutionary events. These banding patterns are particularly useful for finding large scale differences between two genomes or among genomes.Often researchers use bands in studies focused on diseases; to answer the question: do the differences in a genes correlate with a disease?
Karyotype of a typical human male.
However, most of the genetic differences we deal with today are too small
to see with a microscope. These are insertions and deletions of a single
or a few base pairs. Working on this level, other techniques need to be
employed to study DNA. When comparing DNA from one person to another,or
one organism to another, researchers are looking for places where there
are differences or changes from the "ancestral" DNA. The changes
don't have to be repeat differences, they can be the differences in the
distance between the restriction sites or the site itself (a restriction
site is a unique sequence that is recognized by an enzyme).
Normal Strand
of DNA Mutated Strand of DNA
One way to get specific DNA to study is to sort out chromosomes. Flow
sorting employs flow cytometry to separate chromosomes isolated from
cells during cell division when the chromosomes are condensed and stable.As
the chromosomes flow past a laser beam, they are differentiated by analyzing
the amount of DNA present in each, and then the individual chromosomes are
directed to specific collection tubes. Researchers get a full tube of a
particular chromosome. Chromosomes can then be chopped up into sections
of about 30-40,000 nucleotides, using restriction enzymes.
There are two methods used to replicate these pieces of DNA so that researchers
will have enough copies of each to make a thorough study. One is to insert
DNA into E. coli bacteria which will replicate the DNA as the organism
reproduces itself. The second method is Polymerase Chain Reaction.
PCR uses a machine loaded with the chemical building blocks of DNA. Double-stranded
DNA is heat denatured, that is heated until the double strands separate.
Primers in the chemical mixture anneal to the strands and, as the mixture
cools, new DNA is synthesized resulting in a complimentary copy of each
DNA strand. Since primers anneal to each of the single strands of DNA, the
process of synthesizing is very rapid.
One type of DNA sequence that researchers like to study are satellites.
Our genomes are full of repetitive elements. Depending on their length these
repetitions are called satellites, mini-satellites or micro-satellites.
The terms originated during early DNA studies when researchers, purifying
DNA in an ultra-centrifuge, saw bands that had "migrated" to different
places. These bands were called satellites and now this term is applied
to any highly-repetitive element. Studies focus on how these elements originate,
how they vary, and what they do.
Single-point nucleotide polymorphisms (SNPs) are the focus of a lot of investigations
these days. SNPs are more frequent in the genome than other types of polymorphisms,
so that the model of change is a little more straightforward. For instance,
the change from one type of purine to another purine is more common than
a change from a pyrimidine to a purine. Scientists can model these changes
a little bit more accurately when using these kinds of characters.
The big problem for researchers is, when looking at a whole lot of genetic
material, how to rapidly scan for a certain gene or for a difference in
a given gene. A number of methods have been devised to deal with this problem.
Cleavage methods use the enzyme cleavase, which recognizes single-stranded
DNA. The DNA is melted, then rapidly cooled so that it forms a knot. The
cleavase recognizes structures in this knot and gives a fingerprint that
identifies the uniqueness of the particular strand. Another method that
produces a unique structure calls for cleaving the DNA, then heating and
rapidly cooling it, to form a 3-D structure on an electrophoresis gel. Even
a single base pair change will create a different banding pattern on the
gel, because the bands will not migrate at the same rate.
Hetero-duplexes use DNA from two sources - a healthy person and one
with a disease. The DNA region of interest is amplified, then all of the
DNA melted, and allowed to slowly anneal. Some of the "disease"
DNA will join together with the "healthy" DNA and in places where
they are different and the base pairs don't match up, the two strands will
form a bubble or lesion. The protein (DNA repair enzyme) that will bind
the two strands where the mismatch occurs, can be hooked to a magnetic bead
and the entire section can be fished out of solution. The DNA is then re-amplified
to ascertain the specific differences between the the two strands.
Another method of determining differences between DNA is by hybridization.
For instance, a synthetic oligo (a short strand of DNA) is attached to a
sample of DNA. When the strand is heated, a perfect match will melt at a
much higher temperature than an imperfect match and by recording the melting
temperature it can be determined if the sample has the same sequence as
the oligo. Silicon chips coated with synthetic DNA are now beginning to
be used to compare sample DNA.
The Oligo Ligation Assay method analyzes whether or not two DNA sequences are adjacent to one another. If the site where they're found is polymorphic, the oligos will not be ligated. Using two different labeling tags on the two DNA sequences of interest, eg. fluorescent labels and biotin labels, scientists can detect polymorphisms by scoring for the presence or absence of a doubly-tagged ligation product.
The RAPD (random amplified polymorphic DNA) uses oligos that should
"stick" to a number of places in the genome and use them as PCR
primers to generate DNA fragments. Another method used is AFLPs (amplified
fragment length polymorphism). One problem with RAPD's is that the annealing
properties of a short oligo are not desirable, so scientists prefer using
a larger oligo,but larger sequences occur less frequently in the genome.
To solve that they use a restriction enzyme that recognizes a shorter sequence,
cuts it, and then ligate linkers that are longer. Since the short sequence
is known, scientists can use primers to amplify the entire sequence.
One way to organize genetic data and determine genetic relationships is
through the use of trees. The genetic code provides an ideal medium for
computer analysis, as long as the complex data is recorded properly and
the program to analyze the data is designed properly. The DNA sequence already
falls into a code that the computer can read. Once a researcher has characters
that are useful for a certain level of comparison they are fed into the
computer to get an alignment. This alignment is presented in the form of
a tree, much like a family tree. The inherent assumption with that alignment
is the issue of homology. Homology implies that the characters
arose from a common ancestor.
Humans do not form homogenous populations, so genetic differences can be
found between and among groups. When researchers are studying a group of
individuals, they try to find something that's different among the members
of the group, or that differentiates the group from other groups. In this
way the genetic characters are determined to be ancestral. The original
sequence was this, now it's changed to this - so every sequence is a comparison
back to the original sequence. The more we understand about populations
and how they're structured, the more we can apply that structure to the
study of disease.
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Last updated: July 10, 1998