Supernumerary Marker Chromosome Identified in Asian Elephant (Elephas maximus)

Abstract

We identified a small, supernumerary marker chromosome (sSMC) in two phenotypically normal Asian elephants (Elephas maximus): a female (2n = 57,XX,+mar) and her male offspring (2n = 57,XY,+mar). sSMCs are defined as structurally abnormal chromosomes that cannot be identified by conventional banding analysis since they are usually small and often lack distinct banding patterns. Although current molecular techniques can reveal their origin, the mechanism of their formation is not yet fully understood. We determined the origin of the marker using a suite of conventional and molecular cytogenetic approaches that included (a) G- and C-banding, (b) AgNOR staining, (c) preparation of a DNA clone using laser microdissection of the marker chromosome, (d) FISH with commercially available human painting and telomeric probes, and (e) FISH with centromeric DNA derived from the centromeric regions of a marker-free Asian elephant. Moreover, we present new information on the location and number of NORs in Asian and savanna elephants. We show that the metacentric marker was composed of heterochromatin with NORs at the terminal ends, originating most likely from the heterochromatic region of chromosome 27. In this context, we discuss the possible mechanism of marker formation. We also discuss the similarities between sSMCs and B chromosomes and whether the marker chromosome presented here could evolve into a B chromosome in the future.

Keywords: Asian elephant; FISH; NOR; heterochromatin; karyotype; laser microdissection; sSMC; savanna elephant; small supernumerary marker chromosome.

Figure 1

Pedigree chart for the Asian elephant family. Circles represent females and squares represent male individuals. sSMC carriers are marked in black.

Figure 2

G-banded karyotype of Elephas maximus (2n = 57,XX,+mar). The chromosomes were arranged according to Yang et al. [19]. The arrowheads show the NOR positions.

Figure 3

C-banded chromosomes of E. maximus (2n = 57,XX,+mar).

Figure 4

NOR positions in (a) E. maximus (2n = 57,XY,+mar) and (b) L. africana (2n = 56,XX).

Figure 5

(a) FISH of a telomeric probe (green) to the E. maximus, EMA (2n = 57,XY,+mar). (b) The same metaphase spread hybridized with the EMAM1 probe (red). (c) FISH of the LAFM1 probe (red) to the L. africana, LAF chromosomes (2n = 56,XX). The chromosomes are counterstained with DAPI (blue).

Figure 6

Co-hybridization of the EMAM1 (green) and HSA13 probes (red) to metaphase chromosomes of E. maximus (2n = 57,XX,+mar). The HSA13 probe shows signals on the EMA16 chromosome. The chromosomes are counterstained with DAPI (blue).

Figure 7

Schematic reconstruction showing the most possible marker formation during meiosis. The initial break in the (peri)centromeric region of one of the EMA27 homologs led to the horizontal separation of the p- and q-arms producing two isochromosomes, i(27p) and i(27q). One product of the misdivision was the sSMC formed by heterochromatic p-arms, i(27p). The other product was formed by the q-arms of the chromosome, i(27q). During the first meiotic division (MD), the marker along with the normal EMA27 chromosome segregated into one daughter cell. During the second MD, the sister chromatids of the normal EMA27 were released and segregated from one another. The marker chromosome along with one of the normal EMA27 chromatids segregated into the daughter cell, giving rise to an abnormal gamete, and leading to the abnormal zygote with the heterochromatic sSMC after fertilization.